                         SEQUENCE LISTING

<110>  Danisco US, Inc.
       Babe, Lilia
       Goedegebuur, Frits
       Kaper, Thijs
       Pricelius, Sina
 
<120>  Subtilisin Variants and Methods of Use

<130>  NB41644PCT

<150>  62/925265 
<151>  2019-10-24

<150>  62/852337 
<151>  2019-05-24

<160>  7     

<170>  PatentIn version 3.5

<210>  1
<211>  269
<212>  PRT
<213>  Bacillus gibsonii

<400>  1

Gln Gln Thr Val Pro Trp Gly Ile Thr Arg Val Gln Ala Pro Ala Val 
1               5                   10                  15      


His Asn Arg Gly Ile Thr Gly Ser Gly Val Arg Val Ala Ile Leu Asp 
            20                  25                  30          


Ser Gly Ile Ser Ala His Ser Asp Leu Asn Ile Arg Gly Gly Ala Ser 
        35                  40                  45              


Phe Val Pro Gly Glu Pro Thr Thr Ala Asp Leu Asn Gly His Gly Thr 
    50                  55                  60                  


His Val Ala Gly Thr Val Ala Ala Leu Asn Asn Ser Ile Gly Val Ile 
65                  70                  75                  80  


Gly Val Ala Pro Asn Ala Glu Leu Tyr Ala Val Lys Val Leu Gly Ala 
                85                  90                  95      


Asn Gly Ser Gly Ser Val Ser Gly Ile Ala Gln Gly Leu Glu Trp Ala 
            100                 105                 110         


Ala Thr Asn Asn Met His Ile Ala Asn Met Ser Leu Gly Ser Asp Phe 
        115                 120                 125             


Pro Ser Ser Thr Leu Glu Arg Ala Val Asn Tyr Ala Thr Ser Arg Asp 
    130                 135                 140                 


Val Leu Val Ile Ala Ala Thr Gly Asn Asn Gly Ser Gly Ser Val Gly 
145                 150                 155                 160 


Tyr Pro Ala Arg Tyr Ala Asn Ala Met Ala Val Gly Ala Thr Asp Gln 
                165                 170                 175     


Asn Asn Arg Arg Ala Asn Phe Ser Gln Tyr Gly Thr Gly Ile Asp Ile 
            180                 185                 190         


Val Ala Pro Gly Val Asn Val Gln Ser Thr Tyr Pro Gly Asn Arg Tyr 
        195                 200                 205             


Val Ser Met Asn Gly Thr Ser Met Ala Thr Pro His Val Ala Gly Ala 
    210                 215                 220                 


Ala Ala Leu Val Lys Gln Arg Tyr Pro Ser Trp Asn Ala Thr Gln Ile 
225                 230                 235                 240 


Arg Asn His Leu Lys Asn Thr Ala Thr Asn Leu Gly Asn Ser Ser Gln 
                245                 250                 255     


Phe Gly Ser Gly Leu Val Asn Ala Glu Ala Ala Thr Arg 
            260                 265                 


<210>  2
<211>  269
<212>  PRT
<213>  Bacillus gibsonii

<400>  2

Gln Gln Thr Val Pro Trp Gly Ile Thr Arg Val Gln Ala Pro Ala Val 
1               5                   10                  15      


His Asn Arg Gly Ile Thr Gly Ser Gly Val Arg Val Ala Ile Leu Asp 
            20                  25                  30          


Ser Gly Ile Ser Ala His Glu Asp Leu Asn Ile Arg Gly Gly Ala Ser 
        35                  40                  45              


Phe Val Pro Gly Glu Pro Thr Thr Ala Asp Leu Asn Gly His Gly Thr 
    50                  55                  60                  


His Val Ala Gly Thr Val Ala Ala Leu Asn Asn Ser Ile Gly Val Ile 
65                  70                  75                  80  


Gly Val Ala Pro Asn Ala Glu Leu Tyr Ala Val Lys Val Leu Gly Ala 
                85                  90                  95      


Asn Gly Arg Gly Ser Val Ser Gly Ile Ala Gln Gly Leu Glu Trp Ala 
            100                 105                 110         


Ala Thr Asn Asn Met His Ile Ala Asn Met Ser Leu Gly Ala Glu Gly 
        115                 120                 125             


Pro Ser Ser Thr Leu Glu Arg Ala Val Asn Tyr Ala Thr Ser Arg Asp 
    130                 135                 140                 


Val Leu Val Ile Ala Ala Thr Gly Asn Asn Gly Ser Gly Ser Val Gly 
145                 150                 155                 160 


Tyr Pro Ala Arg Tyr Ala Asn Ala Met Ala Val Gly Ala Thr Asp Gln 
                165                 170                 175     


Asn Asn Arg Arg Ala Asn Phe Ser Gln Tyr Gly Thr Gly Ile Asp Ile 
            180                 185                 190         


Val Ala Pro Gly Val Asn Val Gln Ser Thr Tyr Pro Gly Asn Arg Tyr 
        195                 200                 205             


Val Ser Met Asn Gly Thr Ser Met Ala Thr Pro His Val Ala Gly Ala 
    210                 215                 220                 


Ala Ala Leu Val Lys Gln Arg Tyr Pro Ser Trp Asn Ala Thr Gln Ile 
225                 230                 235                 240 


Arg Asn His Leu Lys Asn Thr Ala Thr Asn Leu Gly Asn Ser Ser Gln 
                245                 250                 255     


Phe Gly Ser Gly Leu Val Asn Ala Glu Ala Ala Thr Arg 
            260                 265                 


<210>  3
<211>  980
<212>  DNA
<213>  Bacillus subtilis

<400>  3
ctgacgatat tgcctcctgc ttttccggcc agaccatctt tgtaattaat gccggaataa       60

gcaactttaa tcaggacacc atccttcggc aaatcctctg ttgatatggt tttcacatgg      120

actgaaacat catcggcatt tttttctgcc tgcaaggctt gaaataacgt tgacattcgg      180

cacactcctt ttcatttata tcgtaaccga agaacgttca aaaaaccaaa tcatcaagcc      240

gccattttca cttcgccggc acattgagac aataatggac aaatccggta tcctcttcat      300

agccgttttg ctcatacaag cttcttgcct tccggttgtg gtgctcagtc tgaagtgtta      360

aacattttgc cccgttttgc cctgcataat cctttgcggc agaaagcagc cggccgccgg      420

ctccctttgt acgcgcatga ggaacgacaa ataagtcatt taatatgtat atccttttca      480

ttgacacaga agaaaacgtt ggatagagct gggtaaagcc tatgaattct ccattttctt      540

ctgctatcaa aataacagac tcgtgatttt ccaaacgagc tttcaaaaaa gcctctgccc      600

cttgcaaatc ggatgcctgt ctataaaatt cccgatattg gttaaacagc ggcgcaatgg      660

cggccgcatc tgatgtcttt gcttggcgaa tgttcatctt atttcttcct ccctctcaat      720

aattttttca ttctatccct tttctgtaaa gtttattttt cagaatactt ttatcatcat      780

gctttgaaaa aatatcacga taatatccat tgttctcacg gaagcacacg cgctgataaa      840

cagctgacat caactaaaag tttcattaaa tactttgaaa aaagttgttg acttaaaaga      900

agctaaatgt tatagtaatt gtacagaata gtcttttaag taagtctact ctgaattttt      960

ttaaaaggag agggtaaaga                                                  980


<210>  4
<211>  87
<212>  DNA
<213>  Bacillus subtilis

<400>  4
gtgagaagca aaaaattgtg gatcagcttg ttgtttgcgt taacgttaat ctttacgatg       60

gcgttcagca acatgtctgc gcaggct                                           87


<210>  5
<211>  252
<212>  DNA
<213>  Bacillus lentus

<400>  5
gctgaagaag caaaagaaaa atatttaatt ggctttaatg agcaggaagc tgtcagtgag       60

tttgtagaac aagtagaggc aaatgacgag gtcgccattc tctctgagga agaggaagtc      120

gaaattgaat tgcttcatga atttgaaacg attcctgttt tatccgttga gttaagccca      180

gaagatgtgg acgcgcttga actcgatcca gcgatttctt atattgaaga ggatgcagaa      240

gtaacgacaa tg                                                          252


<210>  6
<211>  247
<212>  DNA
<213>  Bacillus amyloliquefaciens

<400>  6
acataaaaaa ccggccttgg ccccgccggt tttttattat ttttcttcct ccgcatgttc       60

aatccgctcc ataatcgacg gatggctccc tctgaaaatt ttaacgagaa acggcgggtt      120

gacccggctc agtcccgtaa cggccaagtc ctgaaacgtc tcaatcgccg cttcccggtt      180

tccggtcagc tcaatgccgt aacggtcggc ggcgttttcc tgataccggg agacggcatt      240

cgtaatc                                                                247


<210>  7
<211>  2288
<212>  DNA
<213>  artificial sequence

<220>
<223>  Synthesized sequence - 3prime AprE flanking sequence including 
       the kanamycin marker

<400>  7
taggaattag ctgcgatccg cggccgctct agatacataa aaaaccggcc ttggccccgc       60

cggtttttta ttatttttct tcctccgcat gttcaatccg ctccataatc gacggatggc      120

tccctctgaa aattttaacg agaaacggcg ggttgacccg gctcagtccc gtaacggcca      180

agtcctgaaa cgtctcaatc gccgcttccc ggtttccggt cagctcaatg ccgtaacggt      240

cggcggcgtt ttcctgatac cgggagacgg cattcgtaat cagtctttcg actgagcctt      300

tcgttttatt tgatgcctca agctagagag tcattaccag atctcactgc agtcactaaa      360

acaattcatc cagtaaaata taatatttta ttttctccca atcaggcttg atccccagta      420

agtcaaaaaa tagctcgaca tactgttctt ccccgatatc ctccctgatc gaccggacgc      480

agaaggcaat gtcataccac ttgtccgccc tgccgcttct cccaagatca ataaagccac      540

ttactttgcc atctttcaca aagatgttgc tgtctcccag gtcgccgtgg gaaaagacaa      600

gttcctcttc gggcttttcc gtctttaaaa aatcatacag ctcgcgcgga tctttaaatg      660

gagtgtcttc ttcccagttt tcgcaatcca catcggccag atcgttattc agtaagtaat      720

ccaattcggc taagcggctg tctaagctat tcgtataggg acaatccgat atgtcgatgg      780

agtgaaagag cctgatgcac tccgcataca gctcgataat cttttcaggg ctttgttcat      840

cttcatactc ttccgagcaa aggacgccat cggcctcact catgagcaga ttgctccagc      900

catcatgccg ttcaaagtgc aggacctttg gaacaggcag ctttccttcc agccatagca      960

tcatgtcctt ttcccgttcc acatcatagg tggtcccttt ataccggctg tccgtcattt     1020

ttaaatatag gttttcattt tctcccacca gcttatatac cttagcagga gacattcctt     1080

ccgtatcttt tacgcagcgg tatttttcga tcagtttttt caattccggt gatattctca     1140

ttttagccat ttattatttc cttcctcttt tctacagtat ttaaagatac cccaagaagc     1200

taattataac aagacgaact ccaattcact gttccttgca ttctaaaacc ttaaatacca     1260

gaaaacagct ttttcaaagt tgttttgaaa gttggcgtat aacatagtat cgacggagcc     1320

gataacgcct cactcctcac atcaacccgt tacttctatt gtaatcataa attcaaattc     1380

ttagaaccaa gctgtgttcc gcacttttcc acccttttaa gcatggaaac cccgatcgct     1440

gggaaaacta acaatgtttg gagtgatgca aatgaaaaaa atagtggcag ccatcgtggt     1500

aatcggtctt gtgtttatcg cattttttta tctttacagc cgatcaggcg atgtgtatca     1560

atcggtagac gcggatttga tcacactgtc ttcaagcggc caggaagata tcgagattga     1620

aaaaagacag cacgtcaaag atatgctgga tattatgaat cagggaaaac aggtgaagac     1680

agaaaaaaca tcagcccctg attacgaagg gacaatcaag tttcataaag accggtatga     1740

ctcattcaga ctatggattg acggcagcca gcaagccgtt tttttgaagg atggcacata     1800

ctacaaatta agcaaaaatg atacaaaggc gctgctaaat attattaaaa aagaagcaaa     1860

ggattgaaaa tgaaaaagcg aagctaaccg cttcgctttt tcattttatt ggggcaaaat     1920

atctctcagt gcccgtctga gcattttccc cgtcgcattt ttcggaatat cgtcaagaaa     1980

cgtaatggcg gcaggccgct tgtattttgc cagatgcttt tcgcagtgct gcatgatgtc     2040

ctcctctgtt accccagagc gtttcggcac cacatatccc tttaccgctt ccccgctttg     2100

ggggtccggc acgccgatga caaccgcctc cttgacgtcc ggatggctgt acagcacctc     2160

ctccacctcc cgcggataca cattgtatcc tcctacaatg atcatgtctt ttttccggtc     2220

aacaatgtaa aaatagccgt cctcatcccg tcttgccaag tcccccgtat aaagccaccc     2280

gtctttta                                                              2288


