                         SEQUENCE LISTING

<110>  Rational Enzyme Mining REM AB
 
<120>  HUMAN CARBONIC ANHYDRASE II WITH INCREASED PHYSICAL STABILITY

<130>  PC-21063278

<150>  SE 1250400-7
<151>  2012-04-23

<160>  8     

<170>  PatentIn version 3.5

<210>  1
<211>  783
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  Modified human carbonic anhydrase II

<400>  1
atggcccatc actgggggta cggcaaacac aacggacctg agcactggca taaggacttc       60

cccattgcca agggagagcg ccagtcccct gttgacatcg acactcatac agccaagtat      120

gacccttccc tgaagcccct gtctgtttcc tatgatcaag caacttccct gaggatcctc      180

aacaatggtc atgctttcaa cgtggagttt gatgactctc aggacaaagc agtgctcaag      240

ggaggacccc tggatggcac ttacagattg attcagtttc actttcactg gggttcactt      300

gatggacaag gttcagagca tactgtggat aaaaagaaat atgctgcaga acttcacttg      360

gttcactgga acaccaaata tggggatttt gggaaagctg tgcagcaacc tgatggactg      420

gccgttctag gtattttttt gaaggttggc agcgctaaac cgggccttca gaaagttgtt      480

gatgtgctgg attccattaa aacaaagggc aagagtgctg acttcactaa cttcgatcct      540

cgtggcctcc ttcctgaatc cctggattac tggacctacc caggctcact gaccacccct      600

cctcttctgg aatctgtgac ctggattgtg ctcaaggaac ccatcagcgt cagcagcgag      660

caggtgttga aattccgtaa acttaacttc aatggggagg gtgaacccga agaactgatg      720

gtggacaact ggcgcccagc tcagccactg aagaacaggc aaatcaaagc ttccttcaaa      780

taa                                                                    783


<210>  2
<211>  260
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  Modified human carbonic anhydrase II

<400>  2

Met Ser His His Trp Gly Tyr Gly Lys His Asn Gly Pro Glu His Trp 
1               5                   10                  15      


His Lys Asp Phe Pro Ile Ala Lys Gly Glu Arg Gln Ser Pro Val Asp 
            20                  25                  30          


Ile Asp Thr His Thr Ala Lys Tyr Asp Pro Ser Leu Lys Pro Leu Ser 
        35                  40                  45              


Val Ser Tyr Asp Gln Ala Thr Ser Leu Arg Ile Leu Asn Asn Gly His 
    50                  55                  60                  


Ala Phe Asn Val Glu Phe Asp Asp Ser Gln Asp Lys Ala Val Leu Lys 
65                  70                  75                  80  


Gly Gly Pro Leu Asp Gly Thr Tyr Arg Leu Ile Gln Phe His Phe His 
                85                  90                  95      


Trp Gly Ser Leu Asp Gly Gln Gly Ser Glu His Thr Val Asp Lys Lys 
            100                 105                 110         


Lys Tyr Ala Ala Glu Leu His Leu Val His Trp Asn Thr Lys Tyr Gly 
        115                 120                 125             


Asp Phe Gly Lys Ala Val Gln Gln Pro Asp Gly Leu Ala Val Leu Gly 
    130                 135                 140                 


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


Asp Val Leu Asp Ser Ile Lys Thr Lys Gly Lys Ser Ala Asp Phe Thr 
                165                 170                 175     


Asn Phe Asp Pro Arg Gly Leu Leu Pro Glu Ser Leu Asp Tyr Trp Thr 
            180                 185                 190         


Tyr Pro Gly Ser Leu Thr Thr Pro Pro Leu Leu Glu Ser Val Thr Trp 
        195                 200                 205             


Ile Val Leu Lys Glu Pro Ile Ser Val Ser Ser Glu Gln Val Leu Lys 
    210                 215                 220                 


Phe Arg Lys Leu Asn Phe Asn Gly Glu Gly Glu Pro Glu Glu Leu Met 
225                 230                 235                 240 


Val Asp Asn Trp Arg Pro Ala Gln Pro Leu Lys Asn Arg Gln Ile Lys 
                245                 250                 255     


Ala Ser Phe Lys 
            260 


<210>  3
<211>  783
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  Modified human carbonic anhydrase II

<400>  3
atggcccatc actgggggta cggcaaacac aacggacctg agcactggca taaggacttc       60

cccattgcca agggagagcg ccagtcccct gttgacatcg acactcatac agccaagtat      120

gacccttccc tgaagcccct gtctgtttcc tatgatcaag caacttccct gaggatcctc      180

aacaatggtc atgctttcaa cgtggagttt gatgactctc aggacaaagc agtgctcaag      240

ggaggacccc tggatggcac ttacagattg attcagtttc actttcactg gggttgcctt      300

gatggacaag gttcagagca tactgtggat aaaaagaaat atgctgcaga acttcacttg      360

gttcactgga acaccaaata tggggatttt gggaaagctg tgcagcaacc tgatggactg      420

gccgttctag gtattttttt gaaggttggc agcgctaaac cgggccttca gaaagttgtt      480

gatgtgctgg attccattaa aacaaagggc aagagtgctg acttcactaa cttcgatcct      540

cgtggcctcc ttcctgaatc cctggattac tggacctacc caggctcact gaccacccct      600

cctcttctgg aatctgtgac ctggattgtg ctcaaggaac ccatcagcgt cagcagcgag      660

caggtgttga aattccgtaa acttaacttc aatggggagg gtgaacccga agaactgatg      720

tgcgacaact ggcgcccagc tcagccactg aagaacaggc aaatcaaagc ttccttcaaa      780

taa                                                                    783


<210>  4
<211>  260
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  Modified human carbonic anhydrase II


<220>
<221>  BINDING
<222>  (99)..(241)
<223>  Disulfide bridge between aa in position 99 and aa in position 241

<400>  4

Met Ser His His Trp Gly Tyr Gly Lys His Asn Gly Pro Glu His Trp 
1               5                   10                  15      


His Lys Asp Phe Pro Ile Ala Lys Gly Glu Arg Gln Ser Pro Val Asp 
            20                  25                  30          


Ile Asp Thr His Thr Ala Lys Tyr Asp Pro Ser Leu Lys Pro Leu Ser 
        35                  40                  45              


Val Ser Tyr Asp Gln Ala Thr Ser Leu Arg Ile Leu Asn Asn Gly His 
    50                  55                  60                  


Ala Phe Asn Val Glu Phe Asp Asp Ser Gln Asp Lys Ala Val Leu Lys 
65                  70                  75                  80  


Gly Gly Pro Leu Asp Gly Thr Tyr Arg Leu Ile Gln Phe His Phe His 
                85                  90                  95      


Trp Gly Cys Leu Asp Gly Gln Gly Ser Glu His Thr Val Asp Lys Lys 
            100                 105                 110         


Lys Tyr Ala Ala Glu Leu His Leu Val His Trp Asn Thr Lys Tyr Gly 
        115                 120                 125             


Asp Phe Gly Lys Ala Val Gln Gln Pro Asp Gly Leu Ala Val Leu Gly 
    130                 135                 140                 


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


Asp Val Leu Asp Ser Ile Lys Thr Lys Gly Lys Ser Ala Asp Phe Thr 
                165                 170                 175     


Asn Phe Asp Pro Arg Gly Leu Leu Pro Glu Ser Leu Asp Tyr Trp Thr 
            180                 185                 190         


Tyr Pro Gly Ser Leu Thr Thr Pro Pro Leu Leu Glu Ser Val Thr Trp 
        195                 200                 205             


Ile Val Leu Lys Glu Pro Ile Ser Val Ser Ser Glu Gln Val Leu Lys 
    210                 215                 220                 


Phe Arg Lys Leu Asn Phe Asn Gly Glu Gly Glu Pro Glu Glu Leu Met 
225                 230                 235                 240 


Cys Asp Asn Trp Arg Pro Ala Gln Pro Leu Lys Asn Arg Gln Ile Lys 
                245                 250                 255     


Ala Ser Phe Lys 
            260 


<210>  5
<211>  783
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  Modified human carbonic anhydrase II

<400>  5
atggcccatc actgggggta cggcaaacac aacggacctg agcactggca taaggacttc       60

cccatttgca agggagagcg ccagtcccct gttgacatcg acactcatac agccaagtat      120

gacccttccc tgaagcccct gtctgtttcc tatgatcaag caacttccct gaggatcctc      180

aacaatggtc atgctttcaa cgtggagttt gatgactctc aggacaaagc agtgctcaag      240

ggaggacccc tggatggcac ttacagattg attcagtttc actttcactg gggttcactt      300

gatggacaag gttcagagca tactgtggat aaaaagaaat atgctgcaga acttcacttg      360

gttcactgga acaccaaata tggggatttt gggaaagctg tgcagcaacc tgatggactg      420

gccgttctag gtattttttt gaaggttggc agcgctaaac cgggccttca gaaagttgtt      480

gatgtgctgg attccattaa aacaaagggc aagagtgctg acttcactaa cttcgatcct      540

cgtggcctcc ttcctgaatc cctggattac tggacctacc caggctcact gaccacccct      600

ccttgtctgg aatctgtgac ctggattgtg ctcaaggaac ccatcagcgt cagcagcgag      660

caggtgttga aattccgtaa acttaacttc aatggggagg gtgaacccga agaactgatg      720

gtggacaact ggcgcccagc tcagccactg aagaacaggc aaatcaaagc ttccttcaaa      780

taa                                                                    783


<210>  6
<211>  260
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  Modified human carbonic anhydrase II


<220>
<221>  BINDING
<222>  (23)..(202)
<223>  Disulfide bridge between aa in position 23 and aa in position 202

<400>  6

Met Ser His His Trp Gly Tyr Gly Lys His Asn Gly Pro Glu His Trp 
1               5                   10                  15      


His Lys Asp Phe Pro Ile Cys Lys Gly Glu Arg Gln Ser Pro Val Asp 
            20                  25                  30          


Ile Asp Thr His Thr Ala Lys Tyr Asp Pro Ser Leu Lys Pro Leu Ser 
        35                  40                  45              


Val Ser Tyr Asp Gln Ala Thr Ser Leu Arg Ile Leu Asn Asn Gly His 
    50                  55                  60                  


Ala Phe Asn Val Glu Phe Asp Asp Ser Gln Asp Lys Ala Val Leu Lys 
65                  70                  75                  80  


Gly Gly Pro Leu Asp Gly Thr Tyr Arg Leu Ile Gln Phe His Phe His 
                85                  90                  95      


Trp Gly Ser Leu Asp Gly Gln Gly Ser Glu His Thr Val Asp Lys Lys 
            100                 105                 110         


Lys Tyr Ala Ala Glu Leu His Leu Val His Trp Asn Thr Lys Tyr Gly 
        115                 120                 125             


Asp Phe Gly Lys Ala Val Gln Gln Pro Asp Gly Leu Ala Val Leu Gly 
    130                 135                 140                 


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


Asp Val Leu Asp Ser Ile Lys Thr Lys Gly Lys Ser Ala Asp Phe Thr 
                165                 170                 175     


Asn Phe Asp Pro Arg Gly Leu Leu Pro Glu Ser Leu Asp Tyr Trp Thr 
            180                 185                 190         


Tyr Pro Gly Ser Leu Thr Thr Pro Pro Cys Leu Glu Ser Val Thr Trp 
        195                 200                 205             


Ile Val Leu Lys Glu Pro Ile Ser Val Ser Ser Glu Gln Val Leu Lys 
    210                 215                 220                 


Phe Arg Lys Leu Asn Phe Asn Gly Glu Gly Glu Pro Glu Glu Leu Met 
225                 230                 235                 240 


Val Asp Asn Trp Arg Pro Ala Gln Pro Leu Lys Asn Arg Gln Ile Lys 
                245                 250                 255     


Ala Ser Phe Lys 
            260 


<210>  7
<211>  783
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  Modified human carbonic anhydrase II

<400>  7
atggcccatc actgggggta cggcaaacac aacggacctg agcactggca taaggacttc       60

cccatttgca agggagagcg ccagtcccct gttgacatcg acactcatac agccaagtat      120

gacccttccc tgaagcccct gtctgtttcc tatgatcaag caacttccct gaggatcctc      180

aacaatggtc atgctttcaa cgtggagttt gatgactctc aggacaaagc agtgctcaag      240

ggaggacccc tggatggcac ttacagattg attcagtttc actttcactg gggttgcctt      300

gatggacaag gttcagagca tactgtggat aaaaagaaat atgctgcaga acttcacttg      360

gttcactgga acaccaaata tggggatttt gggaaagctg tgcagcaacc tgatggactg      420

gccgttctag gtattttttt gaaggttggc agcgctaaac cgggccttca gaaagttgtt      480

gatgtgctgg attccattaa aacaaagggc aagagtgctg acttcactaa cttcgatcct      540

cgtggcctcc ttcctgaatc cctggattac tggacctacc caggctcact gaccacccct      600

ccttgtctgg aatctgtgac ctggattgtg ctcaaggaac ccatcagcgt cagcagcgag      660

caggtgttga aattccgtaa acttaacttc aatggggagg gtgaacccga agaactgatg      720

tgcgacaact ggcgcccagc tcagccactg aagaacaggc aaatcaaagc ttccttcaaa      780

taa                                                                    783


<210>  8
<211>  260
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  Modified human carbonic anhydrase II


<220>
<221>  BINDING
<222>  (23)..(202)
<223>  Disulfide bridge between aa in position 23 and aa in position 202

<220>
<221>  BINDING
<222>  (99)..(241)
<223>  Disulfide bridge between aa in position 99 and aa in position 241

<400>  8

Met Ser His His Trp Gly Tyr Gly Lys His Asn Gly Pro Glu His Trp 
1               5                   10                  15      


His Lys Asp Phe Pro Ile Cys Lys Gly Glu Arg Gln Ser Pro Val Asp 
            20                  25                  30          


Ile Asp Thr His Thr Ala Lys Tyr Asp Pro Ser Leu Lys Pro Leu Ser 
        35                  40                  45              


Val Ser Tyr Asp Gln Ala Thr Ser Leu Arg Ile Leu Asn Asn Gly His 
    50                  55                  60                  


Ala Phe Asn Val Glu Phe Asp Asp Ser Gln Asp Lys Ala Val Leu Lys 
65                  70                  75                  80  


Gly Gly Pro Leu Asp Gly Thr Tyr Arg Leu Ile Gln Phe His Phe His 
                85                  90                  95      


Trp Gly Cys Leu Asp Gly Gln Gly Ser Glu His Thr Val Asp Lys Lys 
            100                 105                 110         


Lys Tyr Ala Ala Glu Leu His Leu Val His Trp Asn Thr Lys Tyr Gly 
        115                 120                 125             


Asp Phe Gly Lys Ala Val Gln Gln Pro Asp Gly Leu Ala Val Leu Gly 
    130                 135                 140                 


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


Asp Val Leu Asp Ser Ile Lys Thr Lys Gly Lys Ser Ala Asp Phe Thr 
                165                 170                 175     


Asn Phe Asp Pro Arg Gly Leu Leu Pro Glu Ser Leu Asp Tyr Trp Thr 
            180                 185                 190         


Tyr Pro Gly Ser Leu Thr Thr Pro Pro Cys Leu Glu Ser Val Thr Trp 
        195                 200                 205             


Ile Val Leu Lys Glu Pro Ile Ser Val Ser Ser Glu Gln Val Leu Lys 
    210                 215                 220                 


Phe Arg Lys Leu Asn Phe Asn Gly Glu Gly Glu Pro Glu Glu Leu Met 
225                 230                 235                 240 


Cys Asp Asn Trp Arg Pro Ala Gln Pro Leu Lys Asn Arg Gln Ile Lys 
                245                 250                 255     


Ala Ser Phe Lys 
            260 


