                         SEQUENCE LISTING

<110>  Roche Sequencing Solutions, Inc.
 
<120>  LONG LIFETIME ALPHA-HEMOLYSIN NANOPORES

<130>  P33725-WO

<150>  US 62/357,230
<151>  2016-06-30

<160>  22    

<170>  PatentIn version 3.5

<210>  1
<211>  1023
<212>  DNA
<213>  Staphylococcus aureus

<400>  1
atggcagatc tcgatcccgc gaaattaata cgactcacta tagggaggcc acaacggttt       60

ccctctagaa ataattttgt ttaactttaa gaaggagata tacaaatgga ttcagatatt      120

aatattaaaa caggtacaac agatattggt tcaaatacaa cagtaaaaac tggtgattta      180

gtaacttatg ataaagaaaa tggtatgcat aaaaaagtat tttattcttt tattgatgat      240

aaaaatcata ataaaaaatt gttagttatt cgtacaaaag gtactattgc aggtcaatat      300

agagtatata gtgaagaagg tgctaataaa agtggtttag catggccatc tgcttttaaa      360

gttcaattac aattacctga taatgaagta gcacaaattt cagattatta tccacgtaat      420

agtattgata caaaagaata tatgtcaaca ttaacttatg gttttaatgg taatgtaaca      480

ggtgatgata ctggtaaaat tggtggttta attggtgcta atgtttcaat tggtcataca      540

ttaaaatatg tacaaccaga ttttaaaaca attttagaaa gtcctactga taaaaaagtt      600

ggttggaaag taatttttaa taatatggtt aatcaaaatt ggggtcctta tgatcgtgat      660

agttggaatc ctgtatatgg taatcaatta tttatgaaaa caagaaatgg ttctatgaaa      720

gcagctgata atttcttaga tccaaataaa gcatcaagtt tattatcttc aggtttttct      780

cctgattttg caacagttat tactatggat agaaaagcat caaaacaaca aacaaatatt      840

gatgttattt atgaacgtgt aagagatgat tatcaattac attggacatc aactaattgg      900

aaaggtacaa atactaaaga taaatggaca gatagaagtt cagaaagata taaaattgat      960

tgggaaaaag aagaaatgac aaatggtctc agcgcttgga gccacccgca gttcgaaaaa     1020

taa                                                                   1023


<210>  2
<211>  306
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  wild-type alpha-hemolysin with purification tag

<400>  2

Met Ala Asp Ser Asp Ile Asn Ile Lys Thr Gly Thr Thr Asp Ile Gly 
1               5                   10                  15      


Ser Asn Thr Thr Val Lys Thr Gly Asp Leu Val Thr Tyr Asp Lys Glu 
            20                  25                  30          


Asn Gly Met His Lys Lys Val Phe Tyr Ser Phe Ile Asp Asp Lys Asn 
        35                  40                  45              


His Asn Lys Lys Leu Leu Val Ile Arg Thr Lys Gly Thr Ile Ala Gly 
    50                  55                  60                  


Gln Tyr Arg Val Tyr Ser Glu Glu Gly Ala Asn Lys Ser Gly Leu Ala 
65                  70                  75                  80  


Trp Pro Ser Ala Phe Lys Val Gln Leu Gln Leu Pro Asp Asn Glu Val 
                85                  90                  95      


Ala Gln Ile Ser Asp Tyr Tyr Pro Arg Asn Ser Ile Asp Thr Lys Glu 
            100                 105                 110         


Tyr Met Ser Thr Leu Thr Tyr Gly Phe Asn Gly Asn Val Thr Gly Asp 
        115                 120                 125             


Asp Thr Gly Lys Ile Gly Gly Leu Ile Gly Ala Asn Val Ser Ile Gly 
    130                 135                 140                 


His Thr Leu Lys Tyr Val Gln Pro Asp Phe Lys Thr Ile Leu Glu Ser 
145                 150                 155                 160 


Pro Thr Asp Lys Lys Val Gly Trp Lys Val Ile Phe Asn Asn Met Val 
                165                 170                 175     


Asn Gln Asn Trp Gly Pro Tyr Asp Arg Asp Ser Trp Asn Pro Val Tyr 
            180                 185                 190         


Gly Asn Gln Leu Phe Met Lys Thr Arg Asn Gly Ser Met Lys Ala Ala 
        195                 200                 205             


Asp Asn Phe Leu Asp Pro Asn Lys Ala Ser Ser Leu Leu Ser Ser Gly 
    210                 215                 220                 


Phe Ser Pro Asp Phe Ala Thr Val Ile Thr Met Asp Arg Lys Ala Ser 
225                 230                 235                 240 


Lys Gln Gln Thr Asn Ile Asp Val Ile Tyr Glu Arg Val Arg Asp Asp 
                245                 250                 255     


Tyr Gln Leu His Trp Thr Ser Thr Asn Trp Lys Gly Thr Asn Thr Lys 
            260                 265                 270         


Asp Lys Trp Thr Asp Arg Ser Ser Glu Arg Tyr Lys Ile Asp Trp Glu 
        275                 280                 285             


Lys Glu Glu Met Thr Asn Gly Leu Ser Ala Trp Ser His Pro Gln Phe 
    290                 295                 300                 


Glu Lys 
305     


<210>  3
<211>  305
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  mature wild-type alpha-hemolysin with purification tag

<400>  3

Ala Asp Ser Asp Ile Asn Ile Lys Thr Gly Thr Thr Asp Ile Gly Ser 
1               5                   10                  15      


Asn Thr Thr Val Lys Thr Gly Asp Leu Val Thr Tyr Asp Lys Glu Asn 
            20                  25                  30          


Gly Met His Lys Lys Val Phe Tyr Ser Phe Ile Asp Asp Lys Asn His 
        35                  40                  45              


Asn Lys Lys Leu Leu Val Ile Arg Thr Lys Gly Thr Ile Ala Gly Gln 
    50                  55                  60                  


Tyr Arg Val Tyr Ser Glu Glu Gly Ala Asn Lys Ser Gly Leu Ala Trp 
65                  70                  75                  80  


Pro Ser Ala Phe Lys Val Gln Leu Gln Leu Pro Asp Asn Glu Val Ala 
                85                  90                  95      


Gln Ile Ser Asp Tyr Tyr Pro Arg Asn Ser Ile Asp Thr Lys Glu Tyr 
            100                 105                 110         


Met Ser Thr Leu Thr Tyr Gly Phe Asn Gly Asn Val Thr Gly Asp Asp 
        115                 120                 125             


Thr Gly Lys Ile Gly Gly Leu Ile Gly Ala Asn Val Ser Ile Gly His 
    130                 135                 140                 


Thr Leu Lys Tyr Val Gln Pro Asp Phe Lys Thr Ile Leu Glu Ser Pro 
145                 150                 155                 160 


Thr Asp Lys Lys Val Gly Trp Lys Val Ile Phe Asn Asn Met Val Asn 
                165                 170                 175     


Gln Asn Trp Gly Pro Tyr Asp Arg Asp Ser Trp Asn Pro Val Tyr Gly 
            180                 185                 190         


Asn Gln Leu Phe Met Lys Thr Arg Asn Gly Ser Met Lys Ala Ala Asp 
        195                 200                 205             


Asn Phe Leu Asp Pro Asn Lys Ala Ser Ser Leu Leu Ser Ser Gly Phe 
    210                 215                 220                 


Ser Pro Asp Phe Ala Thr Val Ile Thr Met Asp Arg Lys Ala Ser Lys 
225                 230                 235                 240 


Gln Gln Thr Asn Ile Asp Val Ile Tyr Glu Arg Val Arg Asp Asp Tyr 
                245                 250                 255     


Gln Leu His Trp Thr Ser Thr Asn Trp Lys Gly Thr Asn Thr Lys Asp 
            260                 265                 270         


Lys Trp Thr Asp Arg Ser Ser Glu Arg Tyr Lys Ile Asp Trp Glu Lys 
        275                 280                 285             


Glu Glu Met Thr Asn Gly Leu Ser Ala Trp Ser His Pro Gln Phe Glu 
    290                 295                 300                 


Lys 
305 


<210>  4
<211>  293
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  H35G+V149K+H144A alpha-hemolysin

<400>  4

Ala Asp Ser Asp Ile Asn Ile Lys Thr Gly Thr Thr Asp Ile Gly Ser 
1               5                   10                  15      


Asn Thr Thr Val Lys Thr Gly Asp Leu Val Thr Tyr Asp Lys Glu Asn 
            20                  25                  30          


Gly Met Gly Lys Lys Val Phe Tyr Ser Phe Ile Asp Asp Lys Asn His 
        35                  40                  45              


Asn Lys Lys Leu Leu Val Ile Arg Thr Lys Gly Thr Ile Ala Gly Gln 
    50                  55                  60                  


Tyr Arg Val Tyr Ser Glu Glu Gly Ala Asn Lys Ser Gly Leu Ala Trp 
65                  70                  75                  80  


Pro Ser Ala Phe Lys Val Gln Leu Gln Leu Pro Asp Asn Glu Val Ala 
                85                  90                  95      


Gln Ile Ser Asp Tyr Tyr Pro Arg Asn Ser Ile Asp Thr Lys Glu Tyr 
            100                 105                 110         


Met Ser Thr Leu Thr Tyr Gly Phe Asn Gly Asn Val Thr Gly Asp Asp 
        115                 120                 125             


Thr Gly Lys Ile Gly Gly Leu Ile Gly Ala Asn Val Ser Ile Gly Ala 
    130                 135                 140                 


Thr Leu Lys Tyr Lys Gln Pro Asp Phe Lys Thr Ile Leu Glu Ser Pro 
145                 150                 155                 160 


Thr Asp Lys Lys Val Gly Trp Lys Val Ile Phe Asn Asn Met Val Asn 
                165                 170                 175     


Gln Asn Trp Gly Pro Tyr Asp Arg Asp Ser Trp Asn Pro Val Tyr Gly 
            180                 185                 190         


Asn Gln Leu Phe Met Lys Thr Arg Asn Gly Ser Met Lys Ala Ala Asp 
        195                 200                 205             


Asn Phe Leu Asp Pro Asn Lys Ala Ser Ser Leu Leu Ser Ser Gly Phe 
    210                 215                 220                 


Ser Pro Asp Phe Ala Thr Val Ile Thr Met Asp Arg Lys Ala Ser Lys 
225                 230                 235                 240 


Gln Gln Thr Asn Ile Asp Val Ile Tyr Glu Arg Val Arg Asp Asp Tyr 
                245                 250                 255     


Gln Leu His Trp Thr Ser Thr Asn Trp Lys Gly Thr Asn Thr Lys Asp 
            260                 265                 270         


Lys Trp Thr Asp Arg Ser Ser Glu Arg Tyr Lys Ile Asp Trp Glu Lys 
        275                 280                 285             


Glu Glu Met Thr Asn 
    290             


<210>  5
<211>  293
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  H35G+H144A+V149K+E287R alpha-hemolysin

<400>  5

Ala Asp Ser Asp Ile Asn Ile Lys Thr Gly Thr Thr Asp Ile Gly Ser 
1               5                   10                  15      


Asn Thr Thr Val Lys Thr Gly Asp Leu Val Thr Tyr Asp Lys Glu Asn 
            20                  25                  30          


Gly Met Gly Lys Lys Val Phe Tyr Ser Phe Ile Asp Asp Lys Asn His 
        35                  40                  45              


Asn Lys Lys Leu Leu Val Ile Arg Thr Lys Gly Thr Ile Ala Gly Gln 
    50                  55                  60                  


Tyr Arg Val Tyr Ser Glu Glu Gly Ala Asn Lys Ser Gly Leu Ala Trp 
65                  70                  75                  80  


Pro Ser Ala Phe Lys Val Gln Leu Gln Leu Pro Asp Asn Glu Val Ala 
                85                  90                  95      


Gln Ile Ser Asp Tyr Tyr Pro Arg Asn Ser Ile Asp Thr Lys Glu Tyr 
            100                 105                 110         


Met Ser Thr Leu Thr Tyr Gly Phe Asn Gly Asn Val Thr Gly Asp Asp 
        115                 120                 125             


Thr Gly Lys Ile Gly Gly Leu Ile Gly Ala Asn Val Ser Ile Gly Ala 
    130                 135                 140                 


Thr Leu Lys Tyr Lys Gln Pro Asp Phe Lys Thr Ile Leu Glu Ser Pro 
145                 150                 155                 160 


Thr Asp Lys Lys Val Gly Trp Lys Val Ile Phe Asn Asn Met Val Asn 
                165                 170                 175     


Gln Asn Trp Gly Pro Tyr Asp Arg Asp Ser Trp Asn Pro Val Tyr Gly 
            180                 185                 190         


Asn Gln Leu Phe Met Lys Thr Arg Asn Gly Ser Met Lys Ala Ala Asp 
        195                 200                 205             


Asn Phe Leu Asp Pro Asn Lys Ala Ser Ser Leu Leu Ser Ser Gly Phe 
    210                 215                 220                 


Ser Pro Asp Phe Ala Thr Val Ile Thr Met Asp Arg Lys Ala Ser Lys 
225                 230                 235                 240 


Gln Gln Thr Asn Ile Asp Val Ile Tyr Glu Arg Val Arg Asp Asp Tyr 
                245                 250                 255     


Gln Leu His Trp Thr Ser Thr Asn Trp Lys Gly Thr Asn Thr Lys Asp 
            260                 265                 270         


Lys Trp Thr Asp Arg Ser Ser Glu Arg Tyr Lys Ile Asp Trp Arg Lys 
        275                 280                 285             


Glu Glu Met Thr Asn 
    290             


<210>  6
<211>  293
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  V149K+E287R alpha-hemolysin

<400>  6

Ala Asp Ser Asp Ile Asn Ile Lys Thr Gly Thr Thr Asp Ile Gly Ser 
1               5                   10                  15      


Asn Thr Thr Val Lys Thr Gly Asp Leu Val Thr Tyr Asp Lys Glu Asn 
            20                  25                  30          


Gly Met Gly Lys Lys Val Phe Tyr Ser Phe Ile Asp Asp Lys Asn His 
        35                  40                  45              


Asn Lys Lys Leu Leu Val Ile Arg Thr Lys Gly Thr Ile Ala Gly Gln 
    50                  55                  60                  


Tyr Arg Val Tyr Ser Glu Glu Gly Ala Asn Lys Ser Gly Leu Ala Trp 
65                  70                  75                  80  


Pro Ser Ala Phe Lys Val Gln Leu Gln Leu Pro Asp Asn Glu Val Ala 
                85                  90                  95      


Gln Ile Ser Asp Tyr Tyr Pro Arg Asn Ser Ile Asp Thr Lys Glu Tyr 
            100                 105                 110         


Met Ser Thr Leu Thr Tyr Gly Phe Asn Gly Asn Val Thr Gly Asp Asp 
        115                 120                 125             


Thr Gly Lys Ile Gly Gly Leu Ile Gly Ala Asn Val Ser Ile Gly His 
    130                 135                 140                 


Thr Leu Lys Tyr Lys Gln Pro Asp Phe Lys Thr Ile Leu Glu Ser Pro 
145                 150                 155                 160 


Thr Asp Lys Lys Val Gly Trp Lys Val Ile Phe Asn Asn Met Val Asn 
                165                 170                 175     


Gln Asn Trp Gly Pro Tyr Asp Arg Asp Ser Trp Asn Pro Val Tyr Gly 
            180                 185                 190         


Asn Gln Leu Phe Met Lys Thr Arg Asn Gly Ser Met Lys Ala Ala Asp 
        195                 200                 205             


Asn Phe Leu Asp Pro Asn Lys Ala Ser Ser Leu Leu Ser Ser Gly Phe 
    210                 215                 220                 


Ser Pro Asp Phe Ala Thr Val Ile Thr Met Asp Arg Lys Ala Ser Lys 
225                 230                 235                 240 


Gln Gln Thr Asn Ile Asp Val Ile Tyr Glu Arg Val Arg Asp Asp Tyr 
                245                 250                 255     


Gln Leu His Trp Thr Ser Thr Asn Trp Lys Gly Thr Asn Thr Lys Asp 
            260                 265                 270         


Lys Trp Thr Asp Arg Ser Ser Glu Arg Tyr Lys Ile Asp Trp Arg Lys 
        275                 280                 285             


Glu Glu Met Thr Asn 
    290             


<210>  7
<211>  293
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  T109K+H35G alpha-hemolysin

<400>  7

Ala Asp Ser Asp Ile Asn Ile Lys Thr Gly Thr Thr Asp Ile Gly Ser 
1               5                   10                  15      


Asn Thr Thr Val Lys Thr Gly Asp Leu Val Thr Tyr Asp Lys Glu Asn 
            20                  25                  30          


Gly Met Gly Lys Lys Val Phe Tyr Ser Phe Ile Asp Asp Lys Asn His 
        35                  40                  45              


Asn Lys Lys Leu Leu Val Ile Arg Thr Lys Gly Thr Ile Ala Gly Gln 
    50                  55                  60                  


Tyr Arg Val Tyr Ser Glu Glu Gly Ala Asn Lys Ser Gly Leu Ala Trp 
65                  70                  75                  80  


Pro Ser Ala Phe Lys Val Gln Leu Gln Leu Pro Asp Asn Glu Val Ala 
                85                  90                  95      


Gln Ile Ser Asp Tyr Tyr Pro Arg Asn Ser Ile Asp Lys Lys Glu Tyr 
            100                 105                 110         


Met Ser Thr Leu Thr Tyr Gly Phe Asn Gly Asn Val Thr Gly Asp Asp 
        115                 120                 125             


Thr Gly Lys Ile Gly Gly Leu Ile Gly Ala Asn Val Ser Ile Gly Ala 
    130                 135                 140                 


Thr Leu Lys Tyr Val Gln Pro Asp Phe Lys Thr Ile Leu Glu Ser Pro 
145                 150                 155                 160 


Thr Asp Lys Lys Val Gly Trp Lys Val Ile Phe Asn Asn Met Val Asn 
                165                 170                 175     


Gln Asn Trp Gly Pro Tyr Asp Arg Asp Ser Trp Asn Pro Val Tyr Gly 
            180                 185                 190         


Asn Gln Leu Phe Met Lys Thr Arg Asn Gly Ser Met Lys Ala Ala Asp 
        195                 200                 205             


Asn Phe Leu Asp Pro Asn Lys Ala Ser Ser Leu Leu Ser Ser Gly Phe 
    210                 215                 220                 


Ser Pro Asp Phe Ala Thr Val Ile Thr Met Asp Arg Lys Ala Ser Lys 
225                 230                 235                 240 


Gln Gln Thr Asn Ile Asp Val Ile Tyr Glu Arg Val Arg Asp Asp Tyr 
                245                 250                 255     


Gln Leu His Trp Thr Ser Thr Asn Trp Lys Gly Thr Asn Thr Lys Asp 
            260                 265                 270         


Lys Trp Thr Asp Arg Ser Ser Glu Arg Tyr Lys Ile Asp Trp Glu Lys 
        275                 280                 285             


Glu Glu Met Thr Asn 
    290             


<210>  8
<211>  293
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  P151K+H35G alpha-hemolysin

<400>  8

Ala Asp Ser Asp Ile Asn Ile Lys Thr Gly Thr Thr Asp Ile Gly Ser 
1               5                   10                  15      


Asn Thr Thr Val Lys Thr Gly Asp Leu Val Thr Tyr Asp Lys Glu Asn 
            20                  25                  30          


Gly Met Gly Lys Lys Val Phe Tyr Ser Phe Ile Asp Asp Lys Asn His 
        35                  40                  45              


Asn Lys Lys Leu Leu Val Ile Arg Thr Lys Gly Thr Ile Ala Gly Gln 
    50                  55                  60                  


Tyr Arg Val Tyr Ser Glu Glu Gly Ala Asn Lys Ser Gly Leu Ala Trp 
65                  70                  75                  80  


Pro Ser Ala Phe Lys Val Gln Leu Gln Leu Pro Asp Asn Glu Val Ala 
                85                  90                  95      


Gln Ile Ser Asp Tyr Tyr Pro Arg Asn Ser Ile Asp Thr Lys Glu Tyr 
            100                 105                 110         


Met Ser Thr Leu Thr Tyr Gly Phe Asn Gly Asn Val Thr Gly Asp Asp 
        115                 120                 125             


Thr Gly Lys Ile Gly Gly Leu Ile Gly Ala Asn Val Ser Ile Gly Ala 
    130                 135                 140                 


Thr Leu Lys Tyr Val Gln Lys Asp Phe Lys Thr Ile Leu Glu Ser Pro 
145                 150                 155                 160 


Thr Asp Lys Lys Val Gly Trp Lys Val Ile Phe Asn Asn Met Val Asn 
                165                 170                 175     


Gln Asn Trp Gly Pro Tyr Asp Arg Asp Ser Trp Asn Pro Val Tyr Gly 
            180                 185                 190         


Asn Gln Leu Phe Met Lys Thr Arg Asn Gly Ser Met Lys Ala Ala Asp 
        195                 200                 205             


Asn Phe Leu Asp Pro Asn Lys Ala Ser Ser Leu Leu Ser Ser Gly Phe 
    210                 215                 220                 


Ser Pro Asp Phe Ala Thr Val Ile Thr Met Asp Arg Lys Ala Ser Lys 
225                 230                 235                 240 


Gln Gln Thr Asn Ile Asp Val Ile Tyr Glu Arg Val Arg Asp Asp Tyr 
                245                 250                 255     


Gln Leu His Trp Thr Ser Thr Asn Trp Lys Gly Thr Asn Thr Lys Asp 
            260                 265                 270         


Lys Trp Thr Asp Arg Ser Ser Glu Arg Tyr Lys Ile Asp Trp Glu Lys 
        275                 280                 285             


Glu Glu Met Thr Asn 
    290             


<210>  9
<211>  293
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  V149K+P151K+H35G alpha-hemolysin

<400>  9

Ala Asp Ser Asp Ile Asn Ile Lys Thr Gly Thr Thr Asp Ile Gly Ser 
1               5                   10                  15      


Asn Thr Thr Val Lys Thr Gly Asp Leu Val Thr Tyr Asp Lys Glu Asn 
            20                  25                  30          


Gly Met Gly Lys Lys Val Phe Tyr Ser Phe Ile Asp Asp Lys Asn His 
        35                  40                  45              


Asn Lys Lys Leu Leu Val Ile Arg Thr Lys Gly Thr Ile Ala Gly Gln 
    50                  55                  60                  


Tyr Arg Val Tyr Ser Glu Glu Gly Ala Asn Lys Ser Gly Leu Ala Trp 
65                  70                  75                  80  


Pro Ser Ala Phe Lys Val Gln Leu Gln Leu Pro Asp Asn Glu Val Ala 
                85                  90                  95      


Gln Ile Ser Asp Tyr Tyr Pro Arg Asn Ser Ile Asp Thr Lys Glu Tyr 
            100                 105                 110         


Met Ser Thr Leu Thr Tyr Gly Phe Asn Gly Asn Val Thr Gly Asp Asp 
        115                 120                 125             


Thr Gly Lys Ile Gly Gly Leu Ile Gly Ala Asn Val Ser Ile Gly Ala 
    130                 135                 140                 


Thr Leu Lys Tyr Lys Gln Lys Asp Phe Lys Thr Ile Leu Glu Ser Pro 
145                 150                 155                 160 


Thr Asp Lys Lys Val Gly Trp Lys Val Ile Phe Asn Asn Met Val Asn 
                165                 170                 175     


Gln Asn Trp Gly Pro Tyr Asp Arg Asp Ser Trp Asn Pro Val Tyr Gly 
            180                 185                 190         


Asn Gln Leu Phe Met Lys Thr Arg Asn Gly Ser Met Lys Ala Ala Asp 
        195                 200                 205             


Asn Phe Leu Asp Pro Asn Lys Ala Ser Ser Leu Leu Ser Ser Gly Phe 
    210                 215                 220                 


Ser Pro Asp Phe Ala Thr Val Ile Thr Met Asp Arg Lys Ala Ser Lys 
225                 230                 235                 240 


Gln Gln Thr Asn Ile Asp Val Ile Tyr Glu Arg Val Arg Asp Asp Tyr 
                245                 250                 255     


Gln Leu His Trp Thr Ser Thr Asn Trp Lys Gly Thr Asn Thr Lys Asp 
            260                 265                 270         


Lys Trp Thr Asp Arg Ser Ser Glu Arg Tyr Lys Ile Asp Trp Glu Lys 
        275                 280                 285             


Glu Glu Met Thr Asn 
    290             


<210>  10
<211>  293
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  T109K+V149K+H35G

<400>  10

Ala Asp Ser Asp Ile Asn Ile Lys Thr Gly Thr Thr Asp Ile Gly Ser 
1               5                   10                  15      


Asn Thr Thr Val Lys Thr Gly Asp Leu Val Thr Tyr Asp Lys Glu Asn 
            20                  25                  30          


Gly Met Gly Lys Lys Val Phe Tyr Ser Phe Ile Asp Asp Lys Asn His 
        35                  40                  45              


Asn Lys Lys Leu Leu Val Ile Arg Thr Lys Gly Thr Ile Ala Gly Gln 
    50                  55                  60                  


Tyr Arg Val Tyr Ser Glu Glu Gly Ala Asn Lys Ser Gly Leu Ala Trp 
65                  70                  75                  80  


Pro Ser Ala Phe Lys Val Gln Leu Gln Leu Pro Asp Asn Glu Val Ala 
                85                  90                  95      


Gln Ile Ser Asp Tyr Tyr Pro Arg Asn Ser Ile Asp Lys Lys Glu Tyr 
            100                 105                 110         


Met Ser Thr Leu Thr Tyr Gly Phe Asn Gly Asn Val Thr Gly Asp Asp 
        115                 120                 125             


Thr Gly Lys Ile Gly Gly Leu Ile Gly Ala Asn Val Ser Ile Gly Ala 
    130                 135                 140                 


Thr Leu Lys Tyr Lys Gln Pro Asp Phe Lys Thr Ile Leu Glu Ser Pro 
145                 150                 155                 160 


Thr Asp Lys Lys Val Gly Trp Lys Val Ile Phe Asn Asn Met Val Asn 
                165                 170                 175     


Gln Asn Trp Gly Pro Tyr Asp Arg Asp Ser Trp Asn Pro Val Tyr Gly 
            180                 185                 190         


Asn Gln Leu Phe Met Lys Thr Arg Asn Gly Ser Met Lys Ala Ala Asp 
        195                 200                 205             


Asn Phe Leu Asp Pro Asn Lys Ala Ser Ser Leu Leu Ser Ser Gly Phe 
    210                 215                 220                 


Ser Pro Asp Phe Ala Thr Val Ile Thr Met Asp Arg Lys Ala Ser Lys 
225                 230                 235                 240 


Gln Gln Thr Asn Ile Asp Val Ile Tyr Glu Arg Val Arg Asp Asp Tyr 
                245                 250                 255     


Gln Leu His Trp Thr Ser Thr Asn Trp Lys Gly Thr Asn Thr Lys Asp 
            260                 265                 270         


Lys Trp Thr Asp Arg Ser Ser Glu Arg Tyr Lys Ile Asp Trp Glu Lys 
        275                 280                 285             


Glu Glu Met Thr Asn 
    290             


<210>  11
<211>  293
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  V149K+K147N+E111N+127 131G+M113A+H35G alpha-hemolysin

<400>  11

Ala Asp Ser Asp Ile Asn Ile Lys Thr Gly Thr Thr Asp Ile Gly Ser 
1               5                   10                  15      


Asn Thr Thr Val Lys Thr Gly Asp Leu Val Thr Tyr Asp Lys Glu Asn 
            20                  25                  30          


Gly Met Gly Lys Lys Val Phe Tyr Ser Phe Ile Asp Asp Lys Asn His 
        35                  40                  45              


Asn Lys Lys Leu Leu Val Ile Arg Thr Lys Gly Thr Ile Ala Gly Gln 
    50                  55                  60                  


Tyr Arg Val Tyr Ser Glu Glu Gly Ala Asn Lys Ser Gly Leu Ala Trp 
65                  70                  75                  80  


Pro Ser Ala Phe Lys Val Gln Leu Gln Leu Pro Asp Asn Glu Val Ala 
                85                  90                  95      


Gln Ile Ser Asp Tyr Tyr Pro Arg Asn Ser Ile Asp Thr Lys Asn Tyr 
            100                 105                 110         


Ala Ser Thr Leu Thr Tyr Gly Phe Asn Gly Asn Val Thr Gly Gly Gly 
        115                 120                 125             


Gly Gly Gly Ile Gly Gly Leu Ile Gly Ala Asn Val Ser Ile Gly Ala 
    130                 135                 140                 


Thr Leu Asn Tyr Lys Gln Pro Asp Phe Lys Thr Ile Leu Glu Ser Pro 
145                 150                 155                 160 


Thr Asp Lys Lys Val Gly Trp Lys Val Ile Phe Asn Asn Met Val Asn 
                165                 170                 175     


Gln Asn Trp Gly Pro Tyr Asp Arg Asp Ser Trp Asn Pro Val Tyr Gly 
            180                 185                 190         


Asn Gln Leu Phe Met Lys Thr Arg Asn Gly Ser Met Lys Ala Ala Asp 
        195                 200                 205             


Asn Phe Leu Asp Pro Asn Lys Ala Ser Ser Leu Leu Ser Ser Gly Phe 
    210                 215                 220                 


Ser Pro Asp Phe Ala Thr Val Ile Thr Met Asp Arg Lys Ala Ser Lys 
225                 230                 235                 240 


Gln Gln Thr Asn Ile Asp Val Ile Tyr Glu Arg Val Arg Asp Asp Tyr 
                245                 250                 255     


Gln Leu His Trp Thr Ser Thr Asn Trp Lys Gly Thr Asn Thr Lys Asp 
            260                 265                 270         


Lys Trp Thr Asp Arg Ser Ser Glu Arg Tyr Lys Ile Asp Trp Glu Lys 
        275                 280                 285             


Glu Glu Met Thr Asn 
    290             


<210>  12
<211>  293
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  V149K+K147N+E111N+127 131G+M113A alpha-hemolysin

<400>  12

Ala Asp Ser Asp Ile Asn Ile Lys Thr Gly Thr Thr Asp Ile Gly Ser 
1               5                   10                  15      


Asn Thr Thr Val Lys Thr Gly Asp Leu Val Thr Tyr Asp Lys Glu Asn 
            20                  25                  30          


Gly Met His Lys Lys Val Phe Tyr Ser Phe Ile Asp Asp Lys Asn His 
        35                  40                  45              


Asn Lys Lys Leu Leu Val Ile Arg Thr Lys Gly Thr Ile Ala Gly Gln 
    50                  55                  60                  


Tyr Arg Val Tyr Ser Glu Glu Gly Ala Asn Lys Ser Gly Leu Ala Trp 
65                  70                  75                  80  


Pro Ser Ala Phe Lys Val Gln Leu Gln Leu Pro Asp Asn Glu Val Ala 
                85                  90                  95      


Gln Ile Ser Asp Tyr Tyr Pro Arg Asn Ser Ile Asp Thr Lys Asn Tyr 
            100                 105                 110         


Ala Ser Thr Leu Thr Tyr Gly Phe Asn Gly Asn Val Thr Gly Gly Gly 
        115                 120                 125             


Gly Gly Gly Ile Gly Gly Leu Ile Gly Ala Asn Val Ser Ile Gly His 
    130                 135                 140                 


Thr Leu Asn Tyr Lys Gln Pro Asp Phe Lys Thr Ile Leu Glu Ser Pro 
145                 150                 155                 160 


Thr Asp Lys Lys Val Gly Trp Lys Val Ile Phe Asn Asn Met Val Asn 
                165                 170                 175     


Gln Asn Trp Gly Pro Tyr Asp Arg Asp Ser Trp Asn Pro Val Tyr Gly 
            180                 185                 190         


Asn Gln Leu Phe Met Lys Thr Arg Asn Gly Ser Met Lys Ala Ala Asp 
        195                 200                 205             


Asn Phe Leu Asp Pro Asn Lys Ala Ser Ser Leu Leu Ser Ser Gly Phe 
    210                 215                 220                 


Ser Pro Asp Phe Ala Thr Val Ile Thr Met Asp Arg Lys Ala Ser Lys 
225                 230                 235                 240 


Gln Gln Thr Asn Ile Asp Val Ile Tyr Glu Arg Val Arg Asp Asp Tyr 
                245                 250                 255     


Gln Leu His Trp Thr Ser Thr Asn Trp Lys Gly Thr Asn Thr Lys Asp 
            260                 265                 270         


Lys Trp Thr Asp Arg Ser Ser Glu Arg Tyr Lys Ile Asp Trp Glu Lys 
        275                 280                 285             


Glu Glu Met Thr Asn 
    290             


<210>  13
<211>  293
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  T109K+V149K+P151K+H35G alpha-hemolysin

<400>  13

Ala Asp Ser Asp Ile Asn Ile Lys Thr Gly Thr Thr Asp Ile Gly Ser 
1               5                   10                  15      


Asn Thr Thr Val Lys Thr Gly Asp Leu Val Thr Tyr Asp Lys Glu Asn 
            20                  25                  30          


Gly Met Gly Lys Lys Val Phe Tyr Ser Phe Ile Asp Asp Lys Asn His 
        35                  40                  45              


Asn Lys Lys Leu Leu Val Ile Arg Thr Lys Gly Thr Ile Ala Gly Gln 
    50                  55                  60                  


Tyr Arg Val Tyr Ser Glu Glu Gly Ala Asn Lys Ser Gly Leu Ala Trp 
65                  70                  75                  80  


Pro Ser Ala Phe Lys Val Gln Leu Gln Leu Pro Asp Asn Glu Val Ala 
                85                  90                  95      


Gln Ile Ser Asp Tyr Tyr Pro Arg Asn Ser Ile Asp Lys Lys Glu Tyr 
            100                 105                 110         


Met Ser Thr Leu Thr Tyr Gly Phe Asn Gly Asn Val Thr Gly Asp Asp 
        115                 120                 125             


Thr Gly Lys Ile Gly Gly Leu Ile Gly Ala Asn Val Ser Ile Gly Ala 
    130                 135                 140                 


Thr Leu Lys Tyr Lys Gln Lys Asp Phe Lys Thr Ile Leu Glu Ser Pro 
145                 150                 155                 160 


Thr Asp Lys Lys Val Gly Trp Lys Val Ile Phe Asn Asn Met Val Asn 
                165                 170                 175     


Gln Asn Trp Gly Pro Tyr Asp Arg Asp Ser Trp Asn Pro Val Tyr Gly 
            180                 185                 190         


Asn Gln Leu Phe Met Lys Thr Arg Asn Gly Ser Met Lys Ala Ala Asp 
        195                 200                 205             


Asn Phe Leu Asp Pro Asn Lys Ala Ser Ser Leu Leu Ser Ser Gly Phe 
    210                 215                 220                 


Ser Pro Asp Phe Ala Thr Val Ile Thr Met Asp Arg Lys Ala Ser Lys 
225                 230                 235                 240 


Gln Gln Thr Asn Ile Asp Val Ile Tyr Glu Arg Val Arg Asp Asp Tyr 
                245                 250                 255     


Gln Leu His Trp Thr Ser Thr Asn Trp Lys Gly Thr Asn Thr Lys Asp 
            260                 265                 270         


Lys Trp Thr Asp Arg Ser Ser Glu Arg Tyr Lys Ile Asp Trp Glu Lys 
        275                 280                 285             


Glu Glu Met Thr Asn 
    290             


<210>  14
<211>  293
<212>  PRT
<213>  Staphylococcus aureus

<400>  14

Ala Asp Ser Asp Ile Asn Ile Lys Thr Gly Thr Thr Asp Ile Gly Ser 
1               5                   10                  15      


Asn Thr Thr Val Lys Thr Gly Asp Leu Val Thr Tyr Asp Lys Glu Asn 
            20                  25                  30          


Gly Met His Lys Lys Val Phe Tyr Ser Phe Ile Asp Asp Lys Asn His 
        35                  40                  45              


Asn Lys Lys Leu Leu Val Ile Arg Thr Lys Gly Thr Ile Ala Gly Gln 
    50                  55                  60                  


Tyr Arg Val Tyr Ser Glu Glu Gly Ala Asn Lys Ser Gly Leu Ala Trp 
65                  70                  75                  80  


Pro Ser Ala Phe Lys Val Gln Leu Gln Leu Pro Asp Asn Glu Val Ala 
                85                  90                  95      


Gln Ile Ser Asp Tyr Tyr Pro Arg Asn Ser Ile Asp Thr Lys Glu Tyr 
            100                 105                 110         


Met Ser Thr Leu Thr Tyr Gly Phe Asn Gly Asn Val Thr Gly Asp Asp 
        115                 120                 125             


Thr Gly Lys Ile Gly Gly Leu Ile Gly Ala Asn Val Ser Ile Gly His 
    130                 135                 140                 


Thr Leu Lys Tyr Val Gln Pro Asp Phe Lys Thr Ile Leu Glu Ser Pro 
145                 150                 155                 160 


Thr Asp Lys Lys Val Gly Trp Lys Val Ile Phe Asn Asn Met Val Asn 
                165                 170                 175     


Gln Asn Trp Gly Pro Tyr Asp Arg Asp Ser Trp Asn Pro Val Tyr Gly 
            180                 185                 190         


Asn Gln Leu Phe Met Lys Thr Arg Asn Gly Ser Met Lys Ala Ala Asp 
        195                 200                 205             


Asn Phe Leu Asp Pro Asn Lys Ala Ser Ser Leu Leu Ser Ser Gly Phe 
    210                 215                 220                 


Ser Pro Asp Phe Ala Thr Val Ile Thr Met Asp Arg Lys Ala Ser Lys 
225                 230                 235                 240 


Gln Gln Thr Asn Ile Asp Val Ile Tyr Glu Arg Val Arg Asp Asp Tyr 
                245                 250                 255     


Gln Leu His Trp Thr Ser Thr Asn Trp Lys Gly Thr Asn Thr Lys Asp 
            260                 265                 270         


Lys Trp Thr Asp Arg Ser Ser Glu Arg Tyr Lys Ile Asp Trp Glu Lys 
        275                 280                 285             


Glu Glu Met Thr Asn 
    290             


<210>  15
<211>  739
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  His-tagged Clostridium phage phiCPV4 pol6

<400>  15

Met His His His His His His His His Ser Gly Gly Ser Asp Lys His 
1               5                   10                  15      


Thr Gln Tyr Val Lys Glu His Ser Phe Asn Tyr Asp Glu Tyr Lys Lys 
            20                  25                  30          


Ala Asn Phe Asp Lys Ile Glu Cys Leu Ile Phe Asp Thr Glu Ser Cys 
        35                  40                  45              


Thr Asn Tyr Glu Asn Asp Asn Thr Gly Ala Arg Val Tyr Gly Trp Gly 
    50                  55                  60                  


Leu Gly Val Thr Arg Asn His Asn Met Ile Tyr Gly Gln Asn Leu Asn 
65                  70                  75                  80  


Gln Phe Trp Glu Val Cys Gln Asn Ile Phe Asn Asp Trp Tyr His Asp 
                85                  90                  95      


Asn Lys His Thr Ile Lys Ile Thr Lys Thr Lys Lys Gly Phe Pro Lys 
            100                 105                 110         


Arg Lys Tyr Ile Lys Phe Pro Ile Ala Val His Asn Leu Gly Trp Asp 
        115                 120                 125             


Val Glu Phe Leu Lys Tyr Ser Leu Val Glu Asn Gly Phe Asn Tyr Asp 
    130                 135                 140                 


Lys Gly Leu Leu Lys Thr Val Phe Ser Lys Gly Ala Pro Tyr Gln Thr 
145                 150                 155                 160 


Val Thr Asp Val Glu Glu Pro Lys Thr Phe His Ile Val Gln Asn Asn 
                165                 170                 175     


Asn Ile Val Tyr Gly Cys Asn Val Tyr Met Asp Lys Phe Phe Glu Val 
            180                 185                 190         


Glu Asn Lys Asp Gly Ser Thr Thr Glu Ile Gly Leu Cys Leu Asp Phe 
        195                 200                 205             


Phe Asp Ser Tyr Lys Ile Ile Thr Cys Ala Glu Ser Gln Phe His Asn 
    210                 215                 220                 


Tyr Val His Asp Val Asp Pro Met Phe Tyr Lys Met Gly Glu Glu Tyr 
225                 230                 235                 240 


Asp Tyr Asp Thr Trp Arg Ser Pro Thr His Lys Gln Thr Thr Leu Glu 
                245                 250                 255     


Leu Arg Tyr Gln Tyr Asn Asp Ile Tyr Met Leu Arg Glu Val Ile Glu 
            260                 265                 270         


Gln Phe Tyr Ile Asp Gly Leu Cys Gly Gly Glu Leu Pro Leu Thr Gly 
        275                 280                 285             


Met Arg Thr Ala Ser Ser Ile Ala Phe Asn Val Leu Lys Lys Met Thr 
    290                 295                 300                 


Phe Gly Glu Glu Lys Thr Glu Glu Gly Tyr Ile Asn Tyr Phe Glu Leu 
305                 310                 315                 320 


Asp Lys Lys Thr Lys Phe Glu Phe Leu Arg Lys Arg Ile Glu Met Glu 
                325                 330                 335     


Ser Tyr Thr Gly Gly Tyr Thr His Ala Asn His Lys Ala Val Gly Lys 
            340                 345                 350         


Thr Ile Asn Lys Ile Gly Cys Ser Leu Asp Ile Asn Ser Ser Tyr Pro 
        355                 360                 365             


Ser Gln Met Ala Tyr Lys Val Phe Pro Tyr Gly Lys Pro Val Arg Lys 
    370                 375                 380                 


Thr Trp Gly Arg Lys Pro Lys Thr Glu Lys Asn Glu Val Tyr Leu Ile 
385                 390                 395                 400 


Glu Val Gly Phe Asp Phe Val Glu Pro Lys His Glu Glu Tyr Ala Leu 
                405                 410                 415     


Asp Ile Phe Lys Ile Gly Ala Val Asn Ser Lys Ala Leu Ser Pro Ile 
            420                 425                 430         


Thr Gly Ala Val Ser Gly Gln Glu Tyr Phe Cys Thr Asn Ile Lys Asp 
        435                 440                 445             


Gly Lys Ala Ile Pro Val Tyr Lys Glu Leu Lys Asp Thr Lys Leu Thr 
    450                 455                 460                 


Thr Asn Tyr Asn Val Val Leu Thr Ser Val Glu Tyr Glu Phe Trp Ile 
465                 470                 475                 480 


Lys His Phe Asn Phe Gly Val Phe Lys Lys Asp Glu Tyr Asp Cys Phe 
                485                 490                 495     


Glu Val Asp Asn Leu Glu Phe Thr Gly Leu Lys Ile Gly Ser Ile Leu 
            500                 505                 510         


Tyr Tyr Lys Ala Glu Lys Gly Lys Phe Lys Pro Tyr Val Asp His Phe 
        515                 520                 525             


Thr Lys Met Lys Val Glu Asn Lys Lys Leu Gly Asn Lys Pro Leu Thr 
    530                 535                 540                 


Asn Gln Ala Lys Leu Ile Leu Asn Gly Ala Tyr Gly Lys Phe Gly Thr 
545                 550                 555                 560 


Lys Gln Asn Lys Glu Glu Lys Asp Leu Ile Met Asp Lys Asn Gly Leu 
                565                 570                 575     


Leu Thr Phe Thr Gly Ser Val Thr Glu Tyr Glu Gly Lys Glu Phe Tyr 
            580                 585                 590         


Arg Pro Tyr Ala Ser Phe Val Thr Ala Tyr Gly Arg Leu Gln Leu Trp 
        595                 600                 605             


Asn Ala Ile Ile Tyr Ala Val Gly Val Glu Asn Phe Leu Tyr Cys Asp 
    610                 615                 620                 


Thr Asp Ser Ile Tyr Cys Asn Arg Glu Val Asn Ser Leu Ile Glu Asp 
625                 630                 635                 640 


Met Asn Ala Ile Gly Glu Thr Ile Asp Lys Thr Ile Leu Gly Lys Trp 
                645                 650                 655     


Asp Val Glu His Val Phe Asp Lys Phe Lys Val Leu Gly Gln Lys Lys 
            660                 665                 670         


Tyr Met Tyr His Asp Cys Lys Glu Asp Lys Thr Asp Leu Lys Cys Cys 
        675                 680                 685             


Gly Leu Pro Ser Asp Ala Arg Lys Ile Ile Ile Gly Gln Gly Phe Asp 
    690                 695                 700                 


Glu Phe Tyr Leu Gly Lys Asn Val Glu Gly Lys Lys Gln Arg Lys Lys 
705                 710                 715                 720 


Val Ile Gly Gly Cys Leu Leu Leu Asp Thr Leu Phe Thr Ile Lys Lys 
                725                 730                 735     


Ile Met Phe 
            


<210>  16
<211>  17
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  Linker/TEV/HisTag

<400>  16

Gly Leu Ser Ala Glu Asn Leu Tyr Phe Gln Gly His His His His His 
1               5                   10                  15      


His 
    


<210>  17
<211>  293
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  E111N+126-131G+H144A+K147N alpha-hemolysin

<400>  17

Ala Asp Ser Asp Ile Asn Ile Lys Thr Gly Thr Thr Asp Ile Gly Ser 
1               5                   10                  15      


Asn Thr Thr Val Lys Thr Gly Asp Leu Val Thr Tyr Asp Lys Glu Asn 
            20                  25                  30          


Gly Met His Lys Lys Val Phe Tyr Ser Phe Ile Asp Asp Lys Asn His 
        35                  40                  45              


Asn Lys Lys Leu Leu Val Ile Arg Thr Lys Gly Thr Ile Ala Gly Gln 
    50                  55                  60                  


Tyr Arg Val Tyr Ser Glu Glu Gly Ala Asn Lys Ser Gly Leu Ala Trp 
65                  70                  75                  80  


Pro Ser Ala Phe Lys Val Gln Leu Gln Leu Pro Asp Asn Glu Val Ala 
                85                  90                  95      


Gln Ile Ser Asp Tyr Tyr Pro Arg Asn Ser Ile Asp Thr Lys Asn Tyr 
            100                 105                 110         


Met Ser Thr Leu Thr Tyr Gly Phe Asn Gly Asn Val Thr Gly Gly Gly 
        115                 120                 125             


Gly Gly Gly Ile Gly Gly Leu Ile Gly Ala Asn Val Ser Ile Gly Ala 
    130                 135                 140                 


Thr Leu Asn Tyr Val Gln Pro Asp Phe Lys Thr Ile Leu Glu Ser Pro 
145                 150                 155                 160 


Thr Asp Lys Lys Val Gly Trp Lys Val Ile Phe Asn Asn Met Val Asn 
                165                 170                 175     


Gln Asn Trp Gly Pro Tyr Asp Arg Asp Ser Trp Asn Pro Val Tyr Gly 
            180                 185                 190         


Asn Gln Leu Phe Met Lys Thr Arg Asn Gly Ser Met Lys Ala Ala Asp 
        195                 200                 205             


Asn Phe Leu Asp Pro Asn Lys Ala Ser Ser Leu Leu Ser Ser Gly Phe 
    210                 215                 220                 


Ser Pro Asp Phe Ala Thr Val Ile Thr Met Asp Arg Lys Ala Ser Lys 
225                 230                 235                 240 


Gln Gln Thr Asn Ile Asp Val Ile Tyr Glu Arg Val Arg Asp Asp Tyr 
                245                 250                 255     


Gln Leu His Trp Thr Ser Thr Asn Trp Lys Gly Thr Asn Thr Lys Asp 
            260                 265                 270         


Lys Trp Thr Asp Arg Ser Ser Glu Arg Tyr Lys Ile Asp Trp Glu Lys 
        275                 280                 285             


Glu Glu Met Thr Asn 
    290             


<210>  18
<211>  293
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  H35G+E111N+H144A+K147N alpha-hemolysin

<400>  18

Ala Asp Ser Asp Ile Asn Ile Lys Thr Gly Thr Thr Asp Ile Gly Ser 
1               5                   10                  15      


Asn Thr Thr Val Lys Thr Gly Asp Leu Val Thr Tyr Asp Lys Glu Asn 
            20                  25                  30          


Gly Met Gly Lys Lys Val Phe Tyr Ser Phe Ile Asp Asp Lys Asn His 
        35                  40                  45              


Asn Lys Lys Leu Leu Val Ile Arg Thr Lys Gly Thr Ile Ala Gly Gln 
    50                  55                  60                  


Tyr Arg Val Tyr Ser Glu Glu Gly Ala Asn Lys Ser Gly Leu Ala Trp 
65                  70                  75                  80  


Pro Ser Ala Phe Lys Val Gln Leu Gln Leu Pro Asp Asn Glu Val Ala 
                85                  90                  95      


Gln Ile Ser Asp Tyr Tyr Pro Arg Asn Ser Ile Asp Thr Lys Asn Tyr 
            100                 105                 110         


Met Ser Thr Leu Thr Tyr Gly Phe Asn Gly Asn Val Thr Gly Asp Asp 
        115                 120                 125             


Thr Gly Lys Ile Gly Gly Leu Ile Gly Ala Asn Val Ser Ile Gly Ala 
    130                 135                 140                 


Thr Leu Asn Tyr Val Gln Pro Asp Phe Lys Thr Ile Leu Glu Ser Pro 
145                 150                 155                 160 


Thr Asp Lys Lys Val Gly Trp Lys Val Ile Phe Asn Asn Met Val Asn 
                165                 170                 175     


Gln Asn Trp Gly Pro Tyr Asp Arg Asp Ser Trp Asn Pro Val Tyr Gly 
            180                 185                 190         


Asn Gln Leu Phe Met Lys Thr Arg Asn Gly Ser Met Lys Ala Ala Asp 
        195                 200                 205             


Asn Phe Leu Asp Pro Asn Lys Ala Ser Ser Leu Leu Ser Ser Gly Phe 
    210                 215                 220                 


Ser Pro Asp Phe Ala Thr Val Ile Thr Met Asp Arg Lys Ala Ser Lys 
225                 230                 235                 240 


Gln Gln Thr Asn Ile Asp Val Ile Tyr Glu Arg Val Arg Asp Asp Tyr 
                245                 250                 255     


Gln Leu His Trp Thr Ser Thr Asn Trp Lys Gly Thr Asn Thr Lys Asp 
            260                 265                 270         


Lys Trp Thr Asp Arg Ser Ser Glu Arg Tyr Lys Ile Asp Trp Glu Lys 
        275                 280                 285             


Glu Glu Met Thr Asn 
    290             


<210>  19
<211>  293
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  H35G+E111N+M113A+126-131G+H144A+K147N alpha-hemolysin

<400>  19

Ala Asp Ser Asp Ile Asn Ile Lys Thr Gly Thr Thr Asp Ile Gly Ser 
1               5                   10                  15      


Asn Thr Thr Val Lys Thr Gly Asp Leu Val Thr Tyr Asp Lys Glu Asn 
            20                  25                  30          


Gly Met Gly Lys Lys Val Phe Tyr Ser Phe Ile Asp Asp Lys Asn His 
        35                  40                  45              


Asn Lys Lys Leu Leu Val Ile Arg Thr Lys Gly Thr Ile Ala Gly Gln 
    50                  55                  60                  


Tyr Arg Val Tyr Ser Glu Glu Gly Ala Asn Lys Ser Gly Leu Ala Trp 
65                  70                  75                  80  


Pro Ser Ala Phe Lys Val Gln Leu Gln Leu Pro Asp Asn Glu Val Ala 
                85                  90                  95      


Gln Ile Ser Asp Tyr Tyr Pro Arg Asn Ser Ile Asp Thr Lys Asn Tyr 
            100                 105                 110         


Ala Ser Thr Leu Thr Tyr Gly Phe Asn Gly Asn Val Thr Gly Gly Gly 
        115                 120                 125             


Gly Gly Gly Ile Gly Gly Leu Ile Gly Ala Asn Val Ser Ile Gly Ala 
    130                 135                 140                 


Thr Leu Asn Tyr Val Gln Pro Asp Phe Lys Thr Ile Leu Glu Ser Pro 
145                 150                 155                 160 


Thr Asp Lys Lys Val Gly Trp Lys Val Ile Phe Asn Asn Met Val Asn 
                165                 170                 175     


Gln Asn Trp Gly Pro Tyr Asp Arg Asp Ser Trp Asn Pro Val Tyr Gly 
            180                 185                 190         


Asn Gln Leu Phe Met Lys Thr Arg Asn Gly Ser Met Lys Ala Ala Asp 
        195                 200                 205             


Asn Phe Leu Asp Pro Asn Lys Ala Ser Ser Leu Leu Ser Ser Gly Phe 
    210                 215                 220                 


Ser Pro Asp Phe Ala Thr Val Ile Thr Met Asp Arg Lys Ala Ser Lys 
225                 230                 235                 240 


Gln Gln Thr Asn Ile Asp Val Ile Tyr Glu Arg Val Arg Asp Asp Tyr 
                245                 250                 255     


Gln Leu His Trp Thr Ser Thr Asn Trp Lys Gly Thr Asn Thr Lys Asp 
            260                 265                 270         


Lys Trp Thr Asp Arg Ser Ser Glu Arg Tyr Lys Ile Asp Trp Glu Lys 
        275                 280                 285             


Glu Glu Met Thr Asn 
    290             


<210>  20
<211>  293
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  E111N+M113A+126-131G+K147N alpha-hemolysin

<400>  20

Ala Asp Ser Asp Ile Asn Ile Lys Thr Gly Thr Thr Asp Ile Gly Ser 
1               5                   10                  15      


Asn Thr Thr Val Lys Thr Gly Asp Leu Val Thr Tyr Asp Lys Glu Asn 
            20                  25                  30          


Gly Met His Lys Lys Val Phe Tyr Ser Phe Ile Asp Asp Lys Asn His 
        35                  40                  45              


Asn Lys Lys Leu Leu Val Ile Arg Thr Lys Gly Thr Ile Ala Gly Gln 
    50                  55                  60                  


Tyr Arg Val Tyr Ser Glu Glu Gly Ala Asn Lys Ser Gly Leu Ala Trp 
65                  70                  75                  80  


Pro Ser Ala Phe Lys Val Gln Leu Gln Leu Pro Asp Asn Glu Val Ala 
                85                  90                  95      


Gln Ile Ser Asp Tyr Tyr Pro Arg Asn Ser Ile Asp Thr Lys Asn Tyr 
            100                 105                 110         


Ala Ser Thr Leu Thr Tyr Gly Phe Asn Gly Asn Val Thr Gly Gly Gly 
        115                 120                 125             


Gly Gly Gly Ile Gly Gly Leu Ile Gly Ala Asn Val Ser Ile Gly His 
    130                 135                 140                 


Thr Leu Asn Tyr Val Gln Pro Asp Phe Lys Thr Ile Leu Glu Ser Pro 
145                 150                 155                 160 


Thr Asp Lys Lys Val Gly Trp Lys Val Ile Phe Asn Asn Met Val Asn 
                165                 170                 175     


Gln Asn Trp Gly Pro Tyr Asp Arg Asp Ser Trp Asn Pro Val Tyr Gly 
            180                 185                 190         


Asn Gln Leu Phe Met Lys Thr Arg Asn Gly Ser Met Lys Ala Ala Asp 
        195                 200                 205             


Asn Phe Leu Asp Pro Asn Lys Ala Ser Ser Leu Leu Ser Ser Gly Phe 
    210                 215                 220                 


Ser Pro Asp Phe Ala Thr Val Ile Thr Met Asp Arg Lys Ala Ser Lys 
225                 230                 235                 240 


Gln Gln Thr Asn Ile Asp Val Ile Tyr Glu Arg Val Arg Asp Asp Tyr 
                245                 250                 255     


Gln Leu His Trp Thr Ser Thr Asn Trp Lys Gly Thr Asn Thr Lys Asp 
            260                 265                 270         


Lys Trp Thr Asp Arg Ser Ser Glu Arg Tyr Lys Ile Asp Trp Glu Lys 
        275                 280                 285             


Glu Glu Met Thr Asn 
    290             


<210>  21
<211>  23
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  linker-SpyTag protein

<400>  21

Gly Gly Ser Ser Gly Gly Ser Ser Gly Gly Ala His Ile Val Met Val 
1               5                   10                  15      


Asp Ala Tyr Lys Pro Thr Lys 
            20              


<210>  22
<211>  293
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  E111S-M113S-T145S-K147S-L135I alpha-hemolysin

<400>  22

Ala Asp Ser Asp Ile Asn Ile Lys Thr Gly Thr Thr Asp Ile Gly Ser 
1               5                   10                  15      


Asn Thr Thr Val Lys Thr Gly Asp Leu Val Thr Tyr Asp Lys Glu Asn 
            20                  25                  30          


Gly Met His Lys Lys Val Phe Tyr Ser Phe Ile Asp Asp Lys Asn His 
        35                  40                  45              


Asn Lys Lys Leu Leu Val Ile Arg Thr Lys Gly Thr Ile Ala Gly Gln 
    50                  55                  60                  


Tyr Arg Val Tyr Ser Glu Glu Gly Ala Asn Lys Ser Gly Leu Ala Trp 
65                  70                  75                  80  


Pro Ser Ala Phe Lys Val Gln Leu Gln Leu Pro Asp Asn Glu Val Ala 
                85                  90                  95      


Gln Ile Ser Asp Tyr Tyr Pro Arg Asn Ser Ile Asp Thr Lys Ser Tyr 
            100                 105                 110         


Ser Ser Thr Leu Thr Tyr Gly Phe Asn Gly Asn Val Thr Gly Asp Asp 
        115                 120                 125             


Thr Gly Lys Ile Gly Gly Ile Ile Gly Ala Asn Val Ser Ile Gly Ala 
    130                 135                 140                 


Ser Leu Ser Tyr Val Gln Pro Asp Phe Lys Thr Ile Leu Glu Ser Pro 
145                 150                 155                 160 


Thr Asp Lys Lys Val Gly Trp Lys Val Ile Phe Asn Asn Met Val Asn 
                165                 170                 175     


Gln Asn Trp Gly Pro Tyr Asp Arg Asp Ser Trp Asn Pro Val Tyr Gly 
            180                 185                 190         


Asn Gln Leu Phe Met Lys Thr Arg Asn Gly Ser Met Lys Ala Ala Asp 
        195                 200                 205             


Asn Phe Leu Asp Pro Asn Lys Ala Ser Ser Leu Leu Ser Ser Gly Phe 
    210                 215                 220                 


Ser Pro Asp Phe Ala Thr Val Ile Thr Met Asp Arg Lys Ala Ser Lys 
225                 230                 235                 240 


Gln Gln Thr Asn Ile Asp Val Ile Tyr Glu Arg Val Arg Asp Asp Tyr 
                245                 250                 255     


Gln Leu His Trp Thr Ser Thr Asn Trp Lys Gly Thr Asn Thr Lys Asp 
            260                 265                 270         


Lys Trp Thr Asp Arg Ser Ser Glu Arg Tyr Lys Ile Asp Trp Glu Lys 
        275                 280                 285             


Glu Glu Met Thr Asn 
    290             


