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研究生： 史帝芬
Preyesh Stephen
論文名稱： 利用蛋白質工程技術改良澱粉結合蛋白功能之研究
Protein Engineering Methods to Modify the Binding Behavior of Starch Binding Domain (RoCBM21)
指導教授： 呂平江
Lyu, Ping-Chiang
口試委員: 呂平江
Lyu, Ping-Chiang
殷献生
Yin, Hsien-Sheng
詹迺立
Chan, Nei-Li
羅惟正
Lo, Wei-Cheng
張大慈
Chang, Dah-Tsyr
學位類別： 博士
Doctor
系所名稱： 生命科學暨醫學院 - 生物資訊與結構生物研究所
Institute of Bioinformatics and Structural Biology
論文出版年： 2012
畢業學年度： 101
語文別： 英文
論文頁數： 129
中文關鍵詞： 環形序列重組 、澱粉結合區蛋白 、蛋白質工程 、晶體結構
外文關鍵詞： Circular Permutation, Starch Binding Domain Protein, Protein Engineering, Crystal structure
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    • Most carbohydrate degrading enzymes have a two-domain structure consisting of a catalytic domain and a carbohydrate binding module (CBM) domain. CBMs mediate the binding of enzyme to the carbohydrate substrate. Family 21 CBMs contain ~100 amino acid residues, and some members have starch-binding functions or glycogen-binding activities. CBMs have shown to be functional even when it is independent from the enzyme. Glucoamylases containing starch-binding domains (SBDs) are used in a variety of scientific and technological applications. In this dissertation a protein engineering strategy – circular permutation – was employed, which yielded variants with greatly enhanced catalytic performance and modified selectivity. A circularly permutated RoCBM21 (CP90) with improved affinity and selectivity toward longer-chain carbohydrates were synthesized, suggesting a new starch-binding protein may be developed for specific scientific and industrial applications. Many studies using random mutagenesis and DNA shuffling could induce only a minor impact to improve the affinity of SBDs while the circular permutation on RoCBM21 could significantly alter the selectivity and affinity. The thermodynamic and chemical stability of circular permutants were determined as well as the functional characteristics were analyzed. Four of the circular permutants predicted could fold with native-like β-barrel fold. The pH, thermodynamics and chemical stability of the circular permutants were investigated. However significant thermal-denaturation differences were observed in two permutants. The circular permutants expressed diverse rate of binding affinity towards carbohydrates. Circular permutation on the 90th position of amino acid (CP90) could generate a highly efficient candidate with higher binding affinity and higher selectivity towards long chain carbohydrates. Qualitative and quantitative experiments were carried out to substantiate the increased affinity and altered selectivity. Further, we used a standard soluble ligand (amylose EX-I) to characterize the detailed functional and structural aspects of CP90. Site-directed mutagenesis along with the crystal structure reveals an altered binding path, which could be the deciding factor to improve affinity and alter the selectivity of this newly created starch-binding domain. The circularly permuted RoCBM21 (CP90) postulates a novel and potential starch binding domain for efficient industrial applications.

    Abstract………………………………………………...................................... 1 Abbreviations………………………………………….................................... 3 Chapter I. Introduction 1.1. Carbohydrates…………………………………….................................... 4 1.2. Starch and Glycogen……………………………….................................. 5 1.3 β- Cyclodextrin and Amylose EX-I…………………................................ 6 1.4. Rhizopus oryzae………………………………………............................... 7 1.5. Carbohydrate-active Enzymes……………………….............................. 8 1.6. Glucoamylase………………………………………….............................. 9 1.7. Carbohydrate-binding Modules………………………............................ 10 1.8. Starch-binding domain (SBD).………………………….......................... 11 1.9 Applications of Starch Binding Domains (SBD) ……….......................... 13 1.10 Circular permutation of proteins………………………......................... 13 1.11 Functional importance of circular permutation in nature…………… 14 1.12 Circular permutation as a genetic engineering approach……………. 15 1.13 Circular Permutation to modify the functional parameters…………. 16 Tables………………………………….……………………………………… 18 Figures………………………………….…………………………………….. 22 2.0 Objective of the study………………………………….………………… 39 Chapter II. Materials and Methods 2.1. Cloning of Circular Permutated RoCMB21 Genes. ………….………. 40 2.2. Protein Expression and Purification. ………….………………………. 41 2.3 N- Terminal Sequencing………….…………………………….………... 42 2.4 Circular Dichroism (CD) ………….…………………………….………. 42 2.5 Qualitative starch-binding assay………….……………………………... 43 2.6 Quantitative binding assay (Intrinsic Fluorescence)…...….…………… 43 2.7 Isothermal Titration Calorimetry (ITC) ..…….………………………... 44 2.8 Competitive Binding: Fluorescence binding Assay………….…………. 47 2.9 Crystal screening, Molecular Replacement and Refinement.…………. 48 2.10 Analytical ultracentrifugation (AUC) ...……………………………….. 49 2.11 Computational Analysis…..…………………………………………….. 49 2.12 Sequence Details about RoCBM21...…………………………………… 51 Figures…………………………………..…………………………………….. 53 Chapter III. Results 3.1 Construction of Circular Permutatants………………………………… 57 3.2 Characterization of CP90………………………………………………... 58 3.3 Fluorescence Binding Assay……………………………………………... 58 3.4 Amylose EX-I Binding…………………………………..……………….. 59 3.5 Insoluble Starch Binding; Qualitative Binding………………………… 59 3.6 Soluble Starch Binding; Quantitative Binding…………………………. 60 3.7 Competitive Ligand Binding; Fluorescence binding…………………… 60 3.8 Crystal structure of CP90; The dimer interface………………………... 61 3.9 Sedimentation Velocity Analysis; Oligomeric state on ligand binding... 61 3.10 Site-directed mutagenesis; Ligand Binding sites……………………… 62 3.11 Binding Model: Molecular model……………………………………… 63 Tables………………………………………………………………………….. 64 Figures………………………………………………………………………… 69 Chapter IV. Discussion 4.1 Circular permutation…………………………………………………….. 96 4.2 Binding Assays……………………………………………………………. 97 4.3 Crystal structure Analysis; Oligomeric state…………………………… 98 4.4 Ligand Binding sites; Site-directed mutagenesis……………………….. 100 4.5 Proposed alternate binding path; Polysaccharide binding…………….. 101 Tables …………………………………………………………………………. 103 Figures………………………………………………………………………… 104 Chapter V. Conclusions………………………………………………….. 112 Research Publications………………………………………………………... 113 Reference……………………………………………………………………… 114

    1. A. Varki, R. Cummings, J. Esko, H. Freeze, G. Hart, and J. Marth, . Essentials of glycobiology, Cold Spring Harbor Laboratory Press, USA, 1999.
    2. L. Crossman and J. M. Dow, Microbes and infection / Institut Pasteur, 2004, 6, 623-629.
    3. G. S. Kelly, Alternative medicine review : a journal of clinical therapeutic, 1998, 3, 27-39.
    4. K. Gessler, I. Uson, T. Takaha, N. Krauss, S. M. Smith, S. Okada, G. M. Sheldrick and W. Saenger, Proceedings of the National Academy of Sciences of the United States of America, 1999, 96, 4246-4251.
    5. A. M. Smith, K. Denyer and C. R. Martin, Plant physiology, 1995, 107, 673-677.
    6. S. Hizukuri, Shirasaka, K., & Juliano, B., Starch, 1983, 35, 348–350.
    7. S. Hizukuri, Takeda, Y., Yasuda, M., & Suzuki, A., Carbohydrate research, 1983, 94, 205–213.
    8. E. Seif, J. Leigh, Y. Liu, I. Roewer, L. Forget and B. F. Lang, Nucleic acids research, 2005, 33, 734-744.
    9. G. J. Davies and B. Henrissat, Biochemical Society transactions, 2002, 30, 291-297.
    10. B. Henrissat, P. M. Coutinho and G. J. Davies, Plant molecular biology, 2001, 47, 55-72.
    11. E. A. Cameron, M. A. Maynard, C. J. Smith, T. J. Smith, N. M. Koropatkin and E. C. Martens, The Journal of biological chemistry, 2012.
    12. E. Hostinova, A. Solovicova, R. Dvorsky and J. Gasperik, Archives of biochemistry and biophysics, 2003, 411, 189-195.
    13. J. Sevcik, E. Hostinova, A. Solovicova, J. Gasperik, Z. Dauter and K. S. Wilson, The FEBS journal, 2006, 273, 2161-2171.
    14. K. Sorimachi, A. J. Jacks, M. F. Le Gal-Coeffet, G. Williamson, D. B. Archer and M. P. Williamson, Journal of molecular biology, 1996, 259, 970-987.
    15. K. Sorimachi, M. F. Le Gal-Coeffet, G. Williamson, D. B. Archer and M. P. Williamson, Structure, 1997, 5, 647-661.
    16. N. R. Gilkes, R. A. Warren, R. C. Miller, Jr. and D. G. Kilburn, The Journal of biological chemistry, 1988, 263, 10401-10407.
    17. D. N. Bolam, A. Ciruela, S. McQueen-Mason, P. Simpson, M. P. Williamson, J. E. Rixon, A. Boraston, G. P. Hazlewood and H. J. Gilbert, The Biochemical journal, 1998, 331 ( Pt 3), 775-781.
    18. S. M. Southall, P. J. Simpson, H. J. Gilbert, G. Williamson and M. P. Williamson, FEBS letters, 1999, 447, 58-60.
    19. A. L. Carvalho, A. Goyal, J. A. Prates, D. N. Bolam, H. J. Gilbert, V. M. Pires, L. M. Ferreira, A. Planas, M. J. Romao and C. M. Fontes, The Journal of biological chemistry, 2004, 279, 34785-34793.
    20. S. Crennell, E. Garman, G. Laver, E. Vimr and G. Taylor, Structure, 1994, 2, 535-544.
    21. S. Najmudin, C. I. Guerreiro, A. L. Carvalho, J. A. Prates, M. A. Correia, V. D. Alves, L. M. Ferreira, M. J. Romao, H. J. Gilbert, D. N. Bolam and C. M. Fontes, The Journal of biological chemistry, 2006, 281, 8815-8828.
    22. R. B. Tunnicliffe, D. N. Bolam, G. Pell, H. J. Gilbert and M. P. Williamson, Journal of molecular biology, 2005, 347, 287-296.
    23. G. Vaaje-Kolstad, D. R. Houston, A. H. Riemen, V. G. Eijsink and D. M. van Aalten, The Journal of biological chemistry, 2005, 280, 11313-11319.
    24. A. Miyanaga, T. Koseki, H. Matsuzawa, T. Wakagi, H. Shoun and S. Fushinobu, The Journal of biological chemistry, 2004, 279, 44907-44914.
    25. A. B. Boraston, M. Healey, J. Klassen, E. Ficko-Blean, A. Lammerts van Bueren and V. Law, The Journal of biological chemistry, 2006, 281, 587-598.
    26. A. B. Boraston, D. N. Bolam, H. J. Gilbert and G. J. Davies, The Biochemical journal, 2004, 382, 769-781.
    27. W. I. Chou, T. W. Pai, S. H. Liu, B. K. Hsiung and M. D. Chang, The Biochemical journal, 2006, 396, 469-477.
    28. T. Giardina, A. P. Gunning, N. Juge, C. B. Faulds, C. S. Furniss, B. Svensson, V. J. Morris and G. Williamson, Journal of molecular biology, 2001, 313, 1149-1159.
    29. C. Klein and G. E. Schulz, Journal of molecular biology, 1991, 217, 737-750.
    30. C. L. Lawson, R. van Montfort, B. Strokopytov, H. J. Rozeboom, K. H. Kalk, G. E. de Vries, D. Penninga, L. Dijkhuizen and B. W. Dijkstra, Journal of molecular biology, 1994, 236, 590-600.
    31. A. Abe, T. Tonozuka, Y. Sakano and S. Kamitori, Journal of molecular biology, 2004, 335, 811-822.
    32. S. Kamitori, S. Kondo, K. Okuyama, T. Yokota, Y. Shimura, T. Tonozuka and Y. Sakano, Journal of molecular biology, 1999, 287, 907-921.
    33. T. Yokota, T. Tonozuka, Y. Shimura, K. Ichikawa, S. Kamitori and Y. Sakano, Bioscience, biotechnology, and biochemistry, 2001, 65, 619-626.
    34. B. Mikami, H. Iwamoto, D. Malle, H. J. Yoon, E. Demirkan-Sarikaya, Y. Mezaki and Y. Katsuya, Journal of molecular biology, 2006, 359, 690-707.
    35. Y. N. Liu, Y. T. Lai, W. I. Chou, M. D. Chang and P. C. Lyu, The Biochemical journal, 2007, 403, 21-30.
    36. J. Y. Tung, M. D. Chang, W. I. Chou, Y. Y. Liu, Y. H. Yeh, F. Y. Chang, S. C. Lin, Z. L. Qiu and Y. J. Sun, The Biochemical journal, 2008, 416, 27-36.
    37. T. Takahashi, K. Kato, Y. Ikegami and M. Irie, Journal of biochemistry, 1985, 98, 663-671.
    38. P. Dent, A. Lavoinne, S. Nakielny, F. B. Caudwell, P. Watt and P. Cohen, Nature, 1990, 348, 302-308.
    39. J. Xia, S. W. Scherer, P. T. Cohen, M. Majer, T. Xi, R. A. Norman, W. C. Knowler, C. Bogardus and M. Prochazka, Diabetes, 1998, 47, 1519-1524.
    40. C. G. Armstrong, M. J. Doherty and P. T. Cohen, The Biochemical journal, 1998, 336 ( Pt 3), 699-704.
    41. M. Machovic, B. Svensson, E. A. MacGregor and S. Janecek, The FEBS journal, 2005, 272, 5497-5513.
    42. B. K. Dalmia and Z. L. Nikolov, Annals of the New York Academy of Sciences, 1994, 721, 160-167.
    43. Q. Ji, J. P. Vincken, L. C. Suurs and R. G. Visser, Plant molecular biology, 2003, 51, 789-801.
    44. Q. Ji, R. J. Oomen, J. P. Vincken, D. N. Bolam, H. J. Gilbert, L. C. Suurs and R. G. Visser, Plant biotechnology journal, 2004, 2, 251-260.
    45. R. Crittenden, A. Laitila, P. Forssell, J. Matto, M. Saarela, T. Mattila-Sandholm and P. Myllarinen, Applied and environmental microbiology, 2001, 67, 3469-3475.
    46. I. Levy, T. Paldi and O. Shoseyov, Biomaterials, 2004, 25, 1841-1849.
    47. Y. W. Hua, M. C. Chi, H. F. Lo, W. H. Hsu and L. L. Lin, Journal of industrial microbiology & biotechnology, 2004, 31, 273-277.
    48. K. Ohdan, T. Kuriki, H. Takata, H. Kaneko and S. Okada, Applied and environmental microbiology, 2000, 66, 3058-3064.
    49. L. Latorre-Garcia, A. C. Adam, P. Manzanares and J. Polaina, Journal of biotechnology, 2005, 118, 167-176.
    50. N. Juge, J. Nohr, M. F. Le Gal-Coeffet, B. Kramhoft, C. S. Furniss, V. Planchot, D. B. Archer, G. Williamson and B. Svensson, Biochimica et biophysica acta, 2006, 1764, 275-284.
    51. M. T. Fukuyama S, Soong C, Allain E, Viksø- and U. H. Nielsen A, Liu Y, Duan J & Wu W., in Denmark and Novozymes North America, Inc., ed. Novozymes A⁄ S, USA.
    52. S. V. Wang P, Xue H, Johnston DB, Rausch KD and T. ME, Cereal Chem, 2007, 84, 10–14.
    53. T. Pan and O. C. Uhlenbeck, Gene, 1993, 125, 111-114.
    54. J. Weiner, 3rd and E. Bornberg-Bauer, Molecular biology and evolution, 2006, 23, 734-743.
    55. Y. Lindqvist and G. Schneider, Current opinion in structural biology, 1997, 7, 422-427.
    56. S. Uliel, A. Fliess and R. Unger, Protein engineering, 2001, 14, 533-542.
    57. C. P. Ponting and R. B. Russell, Trends in biochemical sciences, 1995, 20, 179-180.
    58. B. A. Cunningham, J. J. Hemperly, T. P. Hopp and G. M. Edelman, Proceedings of the National Academy of Sciences of the United States of America, 1979, 76, 3218-3222.
    59. D. M. Carrington, A. Auffret and D. E. Hanke, Nature, 1985, 313, 64-67.
    60. A. E. Todd, C. A. Orengo and J. M. Thornton, Trends in biochemical sciences, 2002, 27, 419-426.
    61. C. Y. Montanier, M. A. Correia, J. E. Flint, Y. Zhu, A. Basle, L. S. McKee, J. A. Prates, S. J. Polizzi, P. M. Coutinho, R. J. Lewis, B. Henrissat, C. M. Fontes and H. J. Gilbert, The Journal of biological chemistry, 2011, 286, 22499-22509.
    62. D. P. Goldenberg and T. E. Creighton, Journal of molecular biology, 1983, 165, 407-413.
    63. K. Luger, U. Hommel, M. Herold, J. Hofsteenge and K. Kirschner, Science, 1989, 243, 206-210.
    64. T. Zhang, E. Bertelsen, D. Benvegnu and T. Alber, Biochemistry, 1993, 32, 12311-12318.
    65. M. Sagermann, W. A. Baase, B. H. Mooers, L. Gay and B. W. Matthews, Biochemistry, 2004, 43, 1296-1301.
    66. J. M. Abdulhamed, S. al Yousef, M. A. Khan and C. Mullins, British heart journal, 1994, 72, 482-485.
    67. J. L. Johnson and F. M. Raushel, Biochemistry, 1996, 35, 10223-10233.
    68. N. Protasova, M. L. Kireeva, N. V. Murzina, A. G. Murzin, V. N. Uversky, O. I. Gryaznova and A. T. Gudkov, Protein engineering, 1994, 7, 1373-1377.
    69. V. N. Uversky, V. P. Kutyshenko, N. Protasova, V. V. Rogov, K. S. Vassilenko and A. T. Gudkov, Protein science : a publication of the Protein Society, 1996, 5, 1844-1851.
    70. A. K. Svensson, J. A. Zitzewitz, C. R. Matthews and V. F. Smith, Protein engineering, design & selection : PEDS, 2006, 19, 175-185.
    71. S. Topell, J. Hennecke and R. Glockshuber, FEBS letters, 1999, 457, 283-289.
    72. G. S. Baird, D. A. Zacharias and R. Y. Tsien, Proceedings of the National Academy of Sciences of the United States of America, 1999, 96, 11241-11246.
    73. T. Nagai, A. Sawano, E. S. Park and A. Miyawaki, Proceedings of the National Academy of Sciences of the United States of America, 2001, 98, 3197-3202.
    74. K. P. Kent, L. M. Oltrogge and S. G. Boxer, Journal of the American Chemical Society, 2009, 131, 15988-15989.
    75. B. Shui, Q. Wang, F. Lee, L. J. Byrnes, D. M. Chudakov, S. A. Lukyanov, H. Sondermann and M. I. Kotlikoff, PloS one, 2011, 6, e20505.
    76. Y. Li, A. M. Sierra, H. W. Ai and R. E. Campbell, Photochemistry and photobiology, 2008, 84, 111-119.
    77. J. Osuna, A. Perez-Blancas and X. Soberon, Protein engineering, 2002, 15, 463-470.
    78. L. G. Gebhard, V. A. Risso, J. Santos, R. G. Ferreyra, M. E. Noguera and M. R. Ermacora, Journal of molecular biology, 2006, 358, 280-288.
    79. P. Zhang and H. K. Schachman, Protein science : a publication of the Protein Society, 1996, 5, 1290-1300.
    80. V. Chu, S. Freitag, I. Le Trong, R. E. Stenkamp and P. S. Stayton, Protein science : a publication of the Protein Society, 1998, 7, 848-859.
    81. A. V. Cheltsov, M. J. Barber and G. C. Ferreira, The Journal of biological chemistry, 2001, 276, 19141-19149.
    82. E. A. Ribeiro, Jr. and C. H. Ramos, Biochemistry, 2005, 44, 4699-4709.
    83. Z. Qian, J. R. Horton, X. Cheng and S. Lutz, Journal of molecular biology, 2009, 393, 191-201.
    84. K. R. Karukurichi, L. Wang, L. Uzasci, C. M. Manlandro, Q. Wang and P. A. Cole, Journal of the American Chemical Society, 2010, 132, 1222-1223.
    85. Q. Xu, N. D. Rawlings, H. J. Chiu, L. Jaroszewski, H. E. Klock, M. W. Knuth, M. D. Miller, M. A. Elsliger, A. M. Deacon, A. Godzik, S. A. Lesley and I. A. Wilson, PloS one, 2011, 6, e22013.
    86. Y. T. Lee, T. H. Su, W. C. Lo, P. C. Lyu and S. C. Sue, PloS one, 2012, 7, e43820.
    87. T. Nakamura and M. Iwakura, The Journal of biological chemistry, 1999, 274, 19041-19047.
    88. C. Clementi, P. A. Jennings and J. N. Onuchic, Journal of molecular biology, 2001, 311, 879-890.
    89. L. Li and E. I. Shakhnovich, Journal of molecular biology, 2001, 306, 121-132.
    90. E. J. Miller, K. F. Fischer and S. Marqusee, Proceedings of the National Academy of Sciences of the United States of America, 2002, 99, 10359-10363.
    91. M. O. Lindberg, E. Haglund, I. A. Hubner, E. I. Shakhnovich and M. Oliveberg, Proceedings of the National Academy of Sciences of the United States of America, 2006, 103, 4083-4088.
    92. R. J. Kreitman, R. K. Puri and I. Pastan, Proceedings of the National Academy of Sciences of the United States of America, 1994, 91, 6889-6893.
    93. T. U. Schwartz, R. Walczak and G. Blobel, Protein science : a publication of the Protein Society, 2004, 13, 2814-2818.
    94. J. Hennecke, P. Sebbel and R. Glockshuber, J Mol Biol, 1999, 286, 1197-1215.
    95. M. Hahn, K. Piotukh, R. Borriss and U. Heinemann, Proceedings of the National Academy of Sciences of the United States of America, 1994, 91, 10417-10421.
    96. A. Buchwalder, H. Szadkowski and K. Kirschner, Biochemistry, 1992, 31, 1621-1630.
    97. G. Flores, X. Soberon and J. Osuna, Protein science : a publication of the Protein Society, 2004, 13, 1677-1683.
    98. J. S. Butler, D. M. Mitrea, G. Mitrousis, G. Cingolani and S. N. Loh, Biochemistry, 2009, 48, 3497-3507.
    99. S. Reitinger, Y. Yu, J. Wicki, M. Ludwiczek, I. D'Angelo, S. Baturin, M. Okon, N. C. Strynadka, S. Lutz, S. G. Withers and L. P. McIntosh, Biochemistry, 2010, 49, 2464-2474.
    100. Z. Qian and S. Lutz, J Am Chem Soc, 2005, 127, 13466-13467.
    101. Z. Qian, C. J. Fields and S. Lutz, Chembiochem, 2007, 8, 1989-1996.
    102. V. R. Agashe and J. B. Udgaonkar, Biochemistry, 1995, 34, 3286-3299.
    103. Z. O. a. W. Minor, Processing of X-ray Diffraction Data Collected in Oscillation Mode, Academic Press New York, 1997.
    104. M. D. Winn, C. C. Ballard, K. D. Cowtan, E. J. Dodson, P. Emsley, P. R. Evans, R. M. Keegan, E. B. Krissinel, A. G. Leslie, A. McCoy, S. J. McNicholas, G. N. Murshudov, N. S. Pannu, E. A. Potterton, H. R. Powell, R. J. Read, A. Vagin and K. S. Wilson, Acta crystallographica. Section D, Biological crystallography, 2011, 67, 235-242.
    105. A. J. McCoy, R. W. Grosse-Kunstleve, P. D. Adams, M. D. Winn, L. C. Storoni and R. J. Read, Journal of applied crystallography, 2007, 40, 658-674.
    106. P. Emsley and K. Cowtan, Acta crystallographica. Section D, Biological crystallography, 2004, 60, 2126-2132.
    107. A. A. Vagin, R. A. Steiner, A. A. Lebedev, L. Potterton, S. McNicholas, F. Long and G. N. Murshudov, Acta crystallographica. Section D, Biological crystallography, 2004, 60, 2184-2195.
    108. M. M. W. Laskowski R A, Moss D S & Thornton J M J. Appl. Cryst., 1993, 26, 283-291.
    109. W. C. Lo, C. C. Lee, C. Y. Lee and P. C. Lyu, Nucleic acids research, 2009, 37, D328-332.
    110. D. Schneidman-Duhovny, Y. Inbar, R. Nussinov and H. J. Wolfson, Nucleic acids research, 2005, 33, W363-367.
    111. W. C. Lo, C. Y. Lee, C. C. Lee and P. C. Lyu, Nucleic acids research, 2009, 37, W545-551.
    112. M. Wagner, R. Adamczak, A. Porollo and J. Meller, Journal of computational biology : a journal of computational molecular cell biology, 2005, 12, 355-369.
    113. A. C. Wallace, R. A. Laskowski and J. M. Thornton, Protein engineering, 1995, 8, 127-134.
    114. W. C. Lo and P. C. Lyu, Genome Biol, 2008, 9, R11.
    115. Y. Tanaka, K. Tsumoto, M. Umetsu, T. Nakanishi, Y. Yasutake, N. Sakai, M. Yao, I. Tanaka, T. Arakawa and I. Kumagai, Biochemical and biophysical research communications, 2004, 323, 185-191.
    116. U. Haddeland, A. Bennick and F. Brosstad, Thrombosis research, 1995, 77, 329-336.
    117. O. D. Monera, C. M. Kay and R. S. Hodges, Protein science : a publication of the Protein Society, 1994, 3, 1984-1991.
    118. C. N. Pace, Methods in enzymology, 1986, 131, 266-280.
    119. Y. Yu and S. Lutz, Trends Biotechnol, 2011, 29, 18-25.
    120. A. G. J. V. a. D. W. S. W. J.R. Whitaker, Handbook of Food Enzymology, Reilly In 2003.
    121. Candussio, ed. G. E. Schulz, US, 1995, pp. 5, 474, 917.
    122. H. Y. Chang, P. M. Irwin and Z. L. Nikolov, J Biotechnol, 1998, 65, 191-202.
    123. M. F. Le Gal-Coeffet, A. J. Jacks, K. Sorimachi, M. P. Williamson, G. Williamson and D. B. Archer, Eur J Biochem, 1995, 233, 561-567.
    124. E. F. Yue Wang, Roberto da Silva, Allison McDaniel, Janice Seibel, Clark Ford, Starch/Stärke, 2006, 58, 501-508.
    125. M. K. a. M. U. Seizaburo Shiraga, Journal of Molecular Catalysis B: Enzymatic, 2004, 28, 229-234.
    126. B. B. Stoffer, C. Dupont, T. P. Frandsen, J. Lehmbeck and B. Svensson, Protein Eng, 1997, 10, 81-87.
    127. T. Y. Jiang, Y. P. Ci, W. I. Chou, Y. C. Lee, Y. J. Sun, W. Y. Chou, K. M. Li and M. D. Chang, PloS one, 2012, 7, e41131.

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