A structural and kinetic survey of GH5_4 endoglucanases reveals determinants of broad substrate specificity and opportunities for biomass hydrolysis
2020; Elsevier BV; Volume: 295; Issue: 51 Linguagem: Inglês
10.1074/jbc.ra120.015328
ISSN1083-351X
AutoresE.M. Glasgow, Elias I. Kemna, C.A. Bingman, Nicole Ing, Kai Deng, C.M. Bianchetti, Taichi E. Takasuka, Trent R. Northen, Brian G. Fox,
Tópico(s)Microbial Metabolic Engineering and Bioproduction
ResumoBroad-specificity glycoside hydrolases (GHs) contribute to plant biomass hydrolysis by degrading a diverse range of polysaccharides, making them useful catalysts for renewable energy and biocommodity production. Discovery of new GHs with improved kinetic parameters or more tolerant substrate-binding sites could increase the efficiency of renewable bioenergy production even further. GH5 has over 50 subfamilies exhibiting selectivities for reaction with β-(1,4)–linked oligo- and polysaccharides. Among these, subfamily 4 (GH5_4) contains numerous broad-selectivity endoglucanases that hydrolyze cellulose, xyloglucan, and mixed-linkage glucans. We previously surveyed the whole subfamily and found over 100 new broad-specificity endoglucanases, although the structural origins of broad specificity remained unclear. A mechanistic understanding of GH5_4 substrate specificity would help inform the best protein design strategies and the most appropriate industrial application of broad-specificity endoglucanases. Here we report structures of 10 new GH5_4 enzymes from cellulolytic microbes and characterize their substrate selectivity using normalized reducing sugar assays and MS. We found that GH5_4 enzymes have the highest catalytic efficiency for hydrolysis of xyloglucan, glucomannan, and soluble β-glucans, with opportunistic secondary reactions on cellulose, mannan, and xylan. The positions of key aromatic residues determine the overall reaction rate and breadth of substrate tolerance, and they contribute to differences in oligosaccharide cleavage patterns. Our new composite model identifies several critical structural features that confer broad specificity and may be readily engineered into existing industrial enzymes. We demonstrate that GH5_4 endoglucanases can have broad specificity without sacrificing high activity, making them a valuable addition to the biomass deconstruction toolset. Broad-specificity glycoside hydrolases (GHs) contribute to plant biomass hydrolysis by degrading a diverse range of polysaccharides, making them useful catalysts for renewable energy and biocommodity production. Discovery of new GHs with improved kinetic parameters or more tolerant substrate-binding sites could increase the efficiency of renewable bioenergy production even further. GH5 has over 50 subfamilies exhibiting selectivities for reaction with β-(1,4)–linked oligo- and polysaccharides. Among these, subfamily 4 (GH5_4) contains numerous broad-selectivity endoglucanases that hydrolyze cellulose, xyloglucan, and mixed-linkage glucans. We previously surveyed the whole subfamily and found over 100 new broad-specificity endoglucanases, although the structural origins of broad specificity remained unclear. A mechanistic understanding of GH5_4 substrate specificity would help inform the best protein design strategies and the most appropriate industrial application of broad-specificity endoglucanases. Here we report structures of 10 new GH5_4 enzymes from cellulolytic microbes and characterize their substrate selectivity using normalized reducing sugar assays and MS. We found that GH5_4 enzymes have the highest catalytic efficiency for hydrolysis of xyloglucan, glucomannan, and soluble β-glucans, with opportunistic secondary reactions on cellulose, mannan, and xylan. The positions of key aromatic residues determine the overall reaction rate and breadth of substrate tolerance, and they contribute to differences in oligosaccharide cleavage patterns. Our new composite model identifies several critical structural features that confer broad specificity and may be readily engineered into existing industrial enzymes. We demonstrate that GH5_4 endoglucanases can have broad specificity without sacrificing high activity, making them a valuable addition to the biomass deconstruction toolset. Sustainable, biological solutions to the growing climate and energy crises have been the subject of increasing interest in the last 2 decades. In contrast to biofuel production from edible polysaccharides, such as starch, recent efforts have focused on next-generation bioenergy, with higher-energy fuels derived from inedible lignocellulosic biomass found in a variety of abundant plant materials. Enzyme hydrolysis of biomass releases sugars, which can be converted into a growing range of fuels and commodities. Enzymes, however, are a major operational expense in the cellulosic bioenergy process (1Chandel A.K. Garlapati V.K. Singh A.K. Antunes F.A.F. da Silva S.S. The path forward for lignocellulose biorefineries: bottlenecks, solutions, and perspective on commercialization.Bioresour. Technol. 2018; 264 (29960825): 370-38110.1016/j.biortech.2018.06.004Crossref PubMed Scopus (163) Google Scholar), owing to the difficulty of hydrolyzing cellulose relative to starch, prompting search and design efforts for more effective enzyme mixtures. Broad-specificity glycoside hydrolases (GHs) may replace many specialized enzymes with fewer, more flexible catalysts, increasing sugar yield and reducing enzyme variability between feedstocks (2Oke M.A. Annuar M.S.M. Simarani K. Mixed feedstock approach to lignocellulosic ethanol production-prospects and limitations.Bioenerg. Res. 2016; 9: 1189-120310.1007/s12155-016-9765-8Crossref Scopus (41) Google Scholar). One such family rich in broad-specificity cellulases is GH5. GH5 members adopt the (α/β)8-barrel fold and display high structural similarity, despite having low (sometimes <10%) sequence identity (3Nagano N. Orengo C.A. Thornton J.M. One fold with many functions: the evolutionary relationships between TIM barrel families based on their sequences, structures and functions.J. Mol. Biol. 2002; 321 (12206759): 741-76510.1016/s0022-2836(02)00649-6Crossref PubMed Scopus (474) Google Scholar, 4Goldman A.D. Beatty J.T. Landweber L.F. The TIM barrel architecture facilitated the early evolution of protein-mediated metabolism.J. Mol. Evol. 2016; 82 (26733481): 17-2610.1007/s00239-015-9722-8Crossref PubMed Scopus (38) Google Scholar, 5Omelchenko M.V. Galperin M.Y. Wolf Y.I. Koonin E.V. Non-homologous isofunctional enzymes: a systematic analysis of alternative solutions in enzyme evolution.Biol. Direct. 2010; 5 (20433725): 3110.1186/1745-6150-5-31Crossref PubMed Scopus (90) Google Scholar, 6Glasgow E.M. Vander Meulen K.A. Takasuka T.E. Bianchetti C.M. Bergeman L.F. Deutsch S. Fox B.G. Extent and origins of functional diversity in a subfamily of glycoside hydrolases.J. Mol. Biol. 2019; 431 (30685401): 1217-123310.1016/j.jmb.2019.01.024Crossref PubMed Scopus (6) Google Scholar, 7Aspeborg H. Coutinho P.M. Wang Y. Brumer 3rd, H. Henrissat B. Evolution, substrate specificity and subfamily classification of glycoside hydrolase family 5 (GH5).BMC Evol. Biol. 2012; 12 (22992189): 18610.1186/1471-2148-12-186Crossref PubMed Scopus (283) Google Scholar). Variability of function is thought to occur through differences in the loop regions connecting the β-strands to the α-helices, particularly at the C-terminal side of the barrel where the active site is built (8Bianchetti C.M. Brumm P. Smith R.W. Dyer K. Hura G.L. Rutkoski T.J. Phillips Jr., G.N. Structure, dynamics, and specificity of endoglucanase D from Clostridium cellulovorans.J. Mol. Biol. 2013; 425 (23751954): 4267-428510.1016/j.jmb.2013.05.030Crossref PubMed Scopus (27) Google Scholar, 9Meng D.D. Liu X. Dong S. Wang Y.F. Ma X.Q. Zhou H. Wang X. Yao L.S. Feng Y. Li F.L. Structural insights into the substrate specificity of a glycoside hydrolase family 5 lichenase from Caldicellulosiruptor sp. F32.Biochem. J. 2017; 474 (28838949): 3373-338910.1042/BCJ20170328Crossref PubMed Scopus (11) Google Scholar). Functionally relevant structural differences also occur at locations distal to the active site (9Meng D.D. Liu X. Dong S. Wang Y.F. Ma X.Q. Zhou H. Wang X. Yao L.S. Feng Y. Li F.L. Structural insights into the substrate specificity of a glycoside hydrolase family 5 lichenase from Caldicellulosiruptor sp. F32.Biochem. J. 2017; 474 (28838949): 3373-338910.1042/BCJ20170328Crossref PubMed Scopus (11) Google Scholar, 10Attia M.A. Nelson C.E. Offen W.A. Jain N. Davies G.J. Gardner J.G. Brumer H. In vitroin vivo characterization of three Cellvibrio japonicus glycoside hydrolase family 5 members reveals potent xyloglucan backbone-cleaving functions.Biotechnol. Biofuels. 2018; 11 (29467823): 4510.1186/s13068-018-1039-6Crossref PubMed Scopus (15) Google Scholar, 11McGregor N. Morar M. Fenger T.H. Stogios P. Lenfant N. Yin V. Xu X. Evdokimova E. Cui H. Henrissat B. Savchenko A. Brumer H. Structure-function analysis of a mixed-linkage β-glucanase/xyloglucanase from the key ruminal Bacteroidetes Prevotella bryantii B14.J. Biol. Chem. 2016; 291 (26507654): 1175-119710.1074/jbc.M115.691659Abstract Full Text Full Text PDF PubMed Scopus (29) Google Scholar, 12Tseng C.W. Ko T.P. Guo R.T. Huang J.W. Wang H.C. Huang C.H. Cheng Y.S. Wang A.H. Liu J.R. Substrate binding of a GH5 endoglucanase from the ruminal fungus Piromyces rhizinflata.Acta Crystallogr. Sect. F Struct. Biol. Cryst. Commun. 2011; 67 (22102024): 1189-119410.1107/S1744309111032428Crossref PubMed Scopus (21) Google Scholar), where loops may contain disulfide bonds (12Tseng C.W. Ko T.P. Guo R.T. Huang J.W. Wang H.C. Huang C.H. Cheng Y.S. Wang A.H. Liu J.R. Substrate binding of a GH5 endoglucanase from the ruminal fungus Piromyces rhizinflata.Acta Crystallogr. Sect. F Struct. Biol. Cryst. Commun. 2011; 67 (22102024): 1189-119410.1107/S1744309111032428Crossref PubMed Scopus (21) Google Scholar, 13Lo Leggio L. Larsen S. The 1.62 Å structure of Thermoascus aurantiacus endoglucanase: completing the structural picture of subfamilies in glycoside hydrolase family 5.FEBS Lett. 2002; 523 (12123813): 103-10810.1016/s0014-5793(02)02954-xCrossref PubMed Scopus (59) Google Scholar), form short β-sheets or helices (10Attia M.A. Nelson C.E. Offen W.A. Jain N. Davies G.J. Gardner J.G. Brumer H. In vitroin vivo characterization of three Cellvibrio japonicus glycoside hydrolase family 5 members reveals potent xyloglucan backbone-cleaving functions.Biotechnol. Biofuels. 2018; 11 (29467823): 4510.1186/s13068-018-1039-6Crossref PubMed Scopus (15) Google Scholar, 11McGregor N. Morar M. Fenger T.H. Stogios P. Lenfant N. Yin V. Xu X. Evdokimova E. Cui H. Henrissat B. Savchenko A. Brumer H. Structure-function analysis of a mixed-linkage β-glucanase/xyloglucanase from the key ruminal Bacteroidetes Prevotella bryantii B14.J. Biol. Chem. 2016; 291 (26507654): 1175-119710.1074/jbc.M115.691659Abstract Full Text Full Text PDF PubMed Scopus (29) Google Scholar, 12Tseng C.W. Ko T.P. Guo R.T. Huang J.W. Wang H.C. Huang C.H. Cheng Y.S. Wang A.H. Liu J.R. Substrate binding of a GH5 endoglucanase from the ruminal fungus Piromyces rhizinflata.Acta Crystallogr. Sect. F Struct. Biol. Cryst. Commun. 2011; 67 (22102024): 1189-119410.1107/S1744309111032428Crossref PubMed Scopus (21) Google Scholar), or even encompass auxiliary domains (14Badieyan S. Bevan D.R. Zhang C. Study and design of stability in GH5 cellulases.Biotechnol. Bioeng. 2012; 109 (21809329): 31-4410.1002/bit.23280Crossref PubMed Scopus (78) Google Scholar). Because GH5 is one of the largest and most catalytically diverse GH families, a classification scheme was devised that separates the family into over 50 subfamilies based on global analysis of sequences, biochemical data, and structures (7Aspeborg H. Coutinho P.M. Wang Y. Brumer 3rd, H. Henrissat B. Evolution, substrate specificity and subfamily classification of glycoside hydrolase family 5 (GH5).BMC Evol. Biol. 2012; 12 (22992189): 18610.1186/1471-2148-12-186Crossref PubMed Scopus (283) Google Scholar). Subfamily 4 (GH5_4) was noted for being particularly enriched in broad-specificity β-(1,4)-endoglucanases (enzymes that cleave in the middle of long glucan chains, like cellulose). As of writing, the Protein Data Bank (PDB) contains 42 entries for 16 GH5_4 enzymes (Table S1), also summarized in the Carbohydrate-Active Enzyme (CAZy) database (15Lombard V. Golaconda Ramulu H. Drula E. Coutinho P.M. Henrissat B. The carbohydrate-active enzymes database (CAZy) in 2013.Nucleic Acids Res. 2014; 42 (24270786): D490-D49510.1093/nar/gkt1178Crossref PubMed Scopus (3293) Google Scholar), making GH5_4 the second-best structurally characterized subfamily after GH5_2. Many efforts have been directed toward understanding how the sequence and structure of a particular GH5_4 enzyme dictate its substrate selectivity (9Meng D.D. Liu X. Dong S. Wang Y.F. Ma X.Q. Zhou H. Wang X. Yao L.S. Feng Y. Li F.L. Structural insights into the substrate specificity of a glycoside hydrolase family 5 lichenase from Caldicellulosiruptor sp. F32.Biochem. J. 2017; 474 (28838949): 3373-338910.1042/BCJ20170328Crossref PubMed Scopus (11) Google Scholar, 10Attia M.A. Nelson C.E. Offen W.A. Jain N. Davies G.J. Gardner J.G. Brumer H. In vitroin vivo characterization of three Cellvibrio japonicus glycoside hydrolase family 5 members reveals potent xyloglucan backbone-cleaving functions.Biotechnol. Biofuels. 2018; 11 (29467823): 4510.1186/s13068-018-1039-6Crossref PubMed Scopus (15) Google Scholar, 12Tseng C.W. Ko T.P. Guo R.T. Huang J.W. Wang H.C. Huang C.H. Cheng Y.S. Wang A.H. Liu J.R. Substrate binding of a GH5 endoglucanase from the ruminal fungus Piromyces rhizinflata.Acta Crystallogr. Sect. F Struct. Biol. Cryst. Commun. 2011; 67 (22102024): 1189-119410.1107/S1744309111032428Crossref PubMed Scopus (21) Google Scholar, 13Lo Leggio L. Larsen S. The 1.62 Å structure of Thermoascus aurantiacus endoglucanase: completing the structural picture of subfamilies in glycoside hydrolase family 5.FEBS Lett. 2002; 523 (12123813): 103-10810.1016/s0014-5793(02)02954-xCrossref PubMed Scopus (59) Google Scholar, 16Dos Santos C.R. Cordeiro R.L. Wong D.W. Murakami M.T. Structural basis for xyloglucan specificity and α-d-Xylp(1 → 6)-d-Glcp recognition at the −1 subsite within the GH5 family.Biochemistry. 2015; 54 (25714929): 1930-194210.1021/acs.biochem.5b00011Crossref PubMed Scopus (13) Google Scholar, 17Gloster T.M. Ibatullin F.M. Macauley K. Eklöf J.M. Roberts S. Turkenburg J.P. Bjørnvad M.E. Jørgensen P.L. Danielsen S. Johansen K.S. Borchert T.V. Wilson K.S. Brumer H. Davies G.J. Characterization and three-dimensional structures of two distinct bacterial xyloglucanases from families GH5 and GH12.J. Biol. Chem. 2007; 282 (17376777): 19177-1918910.1074/jbc.M700224200Abstract Full Text Full Text PDF PubMed Scopus (86) Google Scholar, 18Venditto I. Najmudin S. Luis A.S. Ferreira L.M. Sakka K. Knox J.P. Gilbert H.J. Fontes C.M. Family 46 carbohydrate-binding modules contribute to the enzymatic hydrolysis of xyloglucan and β-1,3-1,4-glucans through distinct mechanisms.J. Biol. Chem. 2015; 290 (25713075): 10572-1058610.1074/jbc.M115.637827Abstract Full Text Full Text PDF PubMed Scopus (28) Google Scholar, 19Wu T.H. Huang C.H. Ko T.P. Lai H.L. Ma Y. Chen C.C. Cheng Y.S. Liu J.R. Guo R.T. Diverse substrate recognition mechanism revealed by Thermotoga maritima Cel5A structures in complex with cellotetraose, cellobiose and mannotriose.Biochim. Biophys. Acta. 2011; 1814 (21839861): 1832-184010.1016/j.bbapap.2011.07.020Crossref PubMed Scopus (28) Google Scholar). For example, crystal structures with linear (12Tseng C.W. Ko T.P. Guo R.T. Huang J.W. Wang H.C. Huang C.H. Cheng Y.S. Wang A.H. Liu J.R. Substrate binding of a GH5 endoglucanase from the ruminal fungus Piromyces rhizinflata.Acta Crystallogr. Sect. F Struct. Biol. Cryst. Commun. 2011; 67 (22102024): 1189-119410.1107/S1744309111032428Crossref PubMed Scopus (21) Google Scholar, 16Dos Santos C.R. Cordeiro R.L. Wong D.W. Murakami M.T. Structural basis for xyloglucan specificity and α-d-Xylp(1 → 6)-d-Glcp recognition at the −1 subsite within the GH5 family.Biochemistry. 2015; 54 (25714929): 1930-194210.1021/acs.biochem.5b00011Crossref PubMed Scopus (13) Google Scholar, 18Venditto I. Najmudin S. Luis A.S. Ferreira L.M. Sakka K. Knox J.P. Gilbert H.J. Fontes C.M. Family 46 carbohydrate-binding modules contribute to the enzymatic hydrolysis of xyloglucan and β-1,3-1,4-glucans through distinct mechanisms.J. Biol. Chem. 2015; 290 (25713075): 10572-1058610.1074/jbc.M115.637827Abstract Full Text Full Text PDF PubMed Scopus (28) Google Scholar, 19Wu T.H. Huang C.H. Ko T.P. Lai H.L. Ma Y. Chen C.C. Cheng Y.S. Liu J.R. Guo R.T. Diverse substrate recognition mechanism revealed by Thermotoga maritima Cel5A structures in complex with cellotetraose, cellobiose and mannotriose.Biochim. Biophys. Acta. 2011; 1814 (21839861): 1832-184010.1016/j.bbapap.2011.07.020Crossref PubMed Scopus (28) Google Scholar, 20Yuan S.F. Wu T.H. Lee H.L. Hsieh H.Y. Lin W.L. Yang B. Chang C.K. Li Q. Gao J. Huang C.H. Ho M.C. Guo R.T. Liang P.H. Biochemical characterization and structural analysis of a bifunctional cellulase/xylanase from Clostridium thermocellum.J. Biol. Chem. 2015; 290 (25575592): 5739-574810.1074/jbc.M114.604454Abstract Full Text Full Text PDF PubMed Scopus (42) Google Scholar) or branched (9Meng D.D. Liu X. Dong S. Wang Y.F. Ma X.Q. Zhou H. Wang X. Yao L.S. Feng Y. Li F.L. Structural insights into the substrate specificity of a glycoside hydrolase family 5 lichenase from Caldicellulosiruptor sp. F32.Biochem. J. 2017; 474 (28838949): 3373-338910.1042/BCJ20170328Crossref PubMed Scopus (11) Google Scholar, 10Attia M.A. Nelson C.E. Offen W.A. Jain N. Davies G.J. Gardner J.G. Brumer H. In vitroin vivo characterization of three Cellvibrio japonicus glycoside hydrolase family 5 members reveals potent xyloglucan backbone-cleaving functions.Biotechnol. Biofuels. 2018; 11 (29467823): 4510.1186/s13068-018-1039-6Crossref PubMed Scopus (15) Google Scholar, 11McGregor N. Morar M. Fenger T.H. Stogios P. Lenfant N. Yin V. Xu X. Evdokimova E. Cui H. Henrissat B. Savchenko A. Brumer H. Structure-function analysis of a mixed-linkage β-glucanase/xyloglucanase from the key ruminal Bacteroidetes Prevotella bryantii B14.J. Biol. Chem. 2016; 291 (26507654): 1175-119710.1074/jbc.M115.691659Abstract Full Text Full Text PDF PubMed Scopus (29) Google Scholar, 16Dos Santos C.R. Cordeiro R.L. Wong D.W. Murakami M.T. Structural basis for xyloglucan specificity and α-d-Xylp(1 → 6)-d-Glcp recognition at the −1 subsite within the GH5 family.Biochemistry. 2015; 54 (25714929): 1930-194210.1021/acs.biochem.5b00011Crossref PubMed Scopus (13) Google Scholar, 17Gloster T.M. Ibatullin F.M. Macauley K. Eklöf J.M. Roberts S. Turkenburg J.P. Bjørnvad M.E. Jørgensen P.L. Danielsen S. Johansen K.S. Borchert T.V. Wilson K.S. Brumer H. Davies G.J. Characterization and three-dimensional structures of two distinct bacterial xyloglucanases from families GH5 and GH12.J. Biol. Chem. 2007; 282 (17376777): 19177-1918910.1074/jbc.M700224200Abstract Full Text Full Text PDF PubMed Scopus (86) Google Scholar) oligosaccharides bound to the positive (16Dos Santos C.R. Cordeiro R.L. Wong D.W. Murakami M.T. Structural basis for xyloglucan specificity and α-d-Xylp(1 → 6)-d-Glcp recognition at the −1 subsite within the GH5 family.Biochemistry. 2015; 54 (25714929): 1930-194210.1021/acs.biochem.5b00011Crossref PubMed Scopus (13) Google Scholar, 19Wu T.H. Huang C.H. Ko T.P. Lai H.L. Ma Y. Chen C.C. Cheng Y.S. Liu J.R. Guo R.T. Diverse substrate recognition mechanism revealed by Thermotoga maritima Cel5A structures in complex with cellotetraose, cellobiose and mannotriose.Biochim. Biophys. Acta. 2011; 1814 (21839861): 1832-184010.1016/j.bbapap.2011.07.020Crossref PubMed Scopus (28) Google Scholar) or negative (10Attia M.A. Nelson C.E. Offen W.A. Jain N. Davies G.J. Gardner J.G. Brumer H. In vitroin vivo characterization of three Cellvibrio japonicus glycoside hydrolase family 5 members reveals potent xyloglucan backbone-cleaving functions.Biotechnol. Biofuels. 2018; 11 (29467823): 4510.1186/s13068-018-1039-6Crossref PubMed Scopus (15) Google Scholar, 12Tseng C.W. Ko T.P. Guo R.T. Huang J.W. Wang H.C. Huang C.H. Cheng Y.S. Wang A.H. Liu J.R. Substrate binding of a GH5 endoglucanase from the ruminal fungus Piromyces rhizinflata.Acta Crystallogr. Sect. F Struct. Biol. Cryst. Commun. 2011; 67 (22102024): 1189-119410.1107/S1744309111032428Crossref PubMed Scopus (21) Google Scholar, 16Dos Santos C.R. Cordeiro R.L. Wong D.W. Murakami M.T. Structural basis for xyloglucan specificity and α-d-Xylp(1 → 6)-d-Glcp recognition at the −1 subsite within the GH5 family.Biochemistry. 2015; 54 (25714929): 1930-194210.1021/acs.biochem.5b00011Crossref PubMed Scopus (13) Google Scholar, 17Gloster T.M. Ibatullin F.M. Macauley K. Eklöf J.M. Roberts S. Turkenburg J.P. Bjørnvad M.E. Jørgensen P.L. Danielsen S. Johansen K.S. Borchert T.V. Wilson K.S. Brumer H. Davies G.J. Characterization and three-dimensional structures of two distinct bacterial xyloglucanases from families GH5 and GH12.J. Biol. Chem. 2007; 282 (17376777): 19177-1918910.1074/jbc.M700224200Abstract Full Text Full Text PDF PubMed Scopus (86) Google Scholar, 18Venditto I. Najmudin S. Luis A.S. Ferreira L.M. Sakka K. Knox J.P. Gilbert H.J. Fontes C.M. Family 46 carbohydrate-binding modules contribute to the enzymatic hydrolysis of xyloglucan and β-1,3-1,4-glucans through distinct mechanisms.J. Biol. Chem. 2015; 290 (25713075): 10572-1058610.1074/jbc.M115.637827Abstract Full Text Full Text PDF PubMed Scopus (28) Google Scholar, 19Wu T.H. Huang C.H. Ko T.P. Lai H.L. Ma Y. Chen C.C. Cheng Y.S. Liu J.R. Guo R.T. Diverse substrate recognition mechanism revealed by Thermotoga maritima Cel5A structures in complex with cellotetraose, cellobiose and mannotriose.Biochim. Biophys. Acta. 2011; 1814 (21839861): 1832-184010.1016/j.bbapap.2011.07.020Crossref PubMed Scopus (28) Google Scholar, 20Yuan S.F. Wu T.H. Lee H.L. Hsieh H.Y. Lin W.L. Yang B. Chang C.K. Li Q. Gao J. Huang C.H. Ho M.C. Guo R.T. Liang P.H. Biochemical characterization and structural analysis of a bifunctional cellulase/xylanase from Clostridium thermocellum.J. Biol. Chem. 2015; 290 (25575592): 5739-574810.1074/jbc.M114.604454Abstract Full Text Full Text PDF PubMed Scopus (42) Google Scholar) subsites (positive being oriented toward the reducing end of the chain) or spanning the entire cleft (9Meng D.D. Liu X. Dong S. Wang Y.F. Ma X.Q. Zhou H. Wang X. Yao L.S. Feng Y. Li F.L. Structural insights into the substrate specificity of a glycoside hydrolase family 5 lichenase from Caldicellulosiruptor sp. F32.Biochem. J. 2017; 474 (28838949): 3373-338910.1042/BCJ20170328Crossref PubMed Scopus (11) Google Scholar, 11McGregor N. Morar M. Fenger T.H. Stogios P. Lenfant N. Yin V. Xu X. Evdokimova E. Cui H. Henrissat B. Savchenko A. Brumer H. Structure-function analysis of a mixed-linkage β-glucanase/xyloglucanase from the key ruminal Bacteroidetes Prevotella bryantii B14.J. Biol. Chem. 2016; 291 (26507654): 1175-119710.1074/jbc.M115.691659Abstract Full Text Full Text PDF PubMed Scopus (29) Google Scholar) show how sugar chains are coordinated in the active site. Some GH5_4 enzymes prefer highly branched xyloglucan (XG), whereas others hydrolyze only linear polysaccharides (see Fig. S1 for polysaccharide structures), and this has been rationalized by differences in the width and depth of the binding cleft, as well as side pockets to accommodate branched sugars or enforce their positioning (11McGregor N. Morar M. Fenger T.H. Stogios P. Lenfant N. Yin V. Xu X. Evdokimova E. Cui H. Henrissat B. Savchenko A. Brumer H. Structure-function analysis of a mixed-linkage β-glucanase/xyloglucanase from the key ruminal Bacteroidetes Prevotella bryantii B14.J. Biol. Chem. 2016; 291 (26507654): 1175-119710.1074/jbc.M115.691659Abstract Full Text Full Text PDF PubMed Scopus (29) Google Scholar, 12Tseng C.W. Ko T.P. Guo R.T. Huang J.W. Wang H.C. Huang C.H. Cheng Y.S. Wang A.H. Liu J.R. Substrate binding of a GH5 endoglucanase from the ruminal fungus Piromyces rhizinflata.Acta Crystallogr. Sect. F Struct. Biol. Cryst. Commun. 2011; 67 (22102024): 1189-119410.1107/S1744309111032428Crossref PubMed Scopus (21) Google Scholar, 17Gloster T.M. Ibatullin F.M. Macauley K. Eklöf J.M. Roberts S. Turkenburg J.P. Bjørnvad M.E. Jørgensen P.L. Danielsen S. Johansen K.S. Borchert T.V. Wilson K.S. Brumer H. Davies G.J. Characterization and three-dimensional structures of two distinct bacterial xyloglucanases from families GH5 and GH12.J. Biol. Chem. 2007; 282 (17376777): 19177-1918910.1074/jbc.M700224200Abstract Full Text Full Text PDF PubMed Scopus (86) Google Scholar, 18Venditto I. Najmudin S. Luis A.S. Ferreira L.M. Sakka K. Knox J.P. Gilbert H.J. Fontes C.M. Family 46 carbohydrate-binding modules contribute to the enzymatic hydrolysis of xyloglucan and β-1,3-1,4-glucans through distinct mechanisms.J. Biol. Chem. 2015; 290 (25713075): 10572-1058610.1074/jbc.M115.637827Abstract Full Text Full Text PDF PubMed Scopus (28) Google Scholar, 20Yuan S.F. Wu T.H. Lee H.L. Hsieh H.Y. Lin W.L. Yang B. Chang C.K. Li Q. Gao J. Huang C.H. Ho M.C. Guo R.T. Liang P.H. Biochemical characterization and structural analysis of a bifunctional cellulase/xylanase from Clostridium thermocellum.J. Biol. Chem. 2015; 290 (25575592): 5739-574810.1074/jbc.M114.604454Abstract Full Text Full Text PDF PubMed Scopus (42) Google Scholar). The ability of some enzymes (6Glasgow E.M. Vander Meulen K.A. Takasuka T.E. Bianchetti C.M. Bergeman L.F. Deutsch S. Fox B.G. Extent and origins of functional diversity in a subfamily of glycoside hydrolases.J. Mol. Biol. 2019; 431 (30685401): 1217-123310.1016/j.jmb.2019.01.024Crossref PubMed Scopus (6) Google Scholar, 7Aspeborg H. Coutinho P.M. Wang Y. Brumer 3rd, H. Henrissat B. Evolution, substrate specificity and subfamily classification of glycoside hydrolase family 5 (GH5).BMC Evol. Biol. 2012; 12 (22992189): 18610.1186/1471-2148-12-186Crossref PubMed Scopus (283) Google Scholar, 11McGregor N. Morar M. Fenger T.H. Stogios P. Lenfant N. Yin V. Xu X. Evdokimova E. Cui H. Henrissat B. Savchenko A. Brumer H. Structure-function analysis of a mixed-linkage β-glucanase/xyloglucanase from the key ruminal Bacteroidetes Prevotella bryantii B14.J. Biol. Chem. 2016; 291 (26507654): 1175-119710.1074/jbc.M115.691659Abstract Full Text Full Text PDF PubMed Scopus (29) Google Scholar, 18Venditto I. Najmudin S. Luis A.S. Ferreira L.M. Sakka K. Knox J.P. Gilbert H.J. Fontes C.M. Family 46 carbohydrate-binding modules contribute to the enzymatic hydrolysis of xyloglucan and β-1,3-1,4-glucans through distinct mechanisms.J. Biol. Chem. 2015; 290 (25713075): 10572-1058610.1074/jbc.M115.637827Abstract Full Text Full Text PDF PubMed Scopus (28) Google Scholar) to hydrolyze both β-(1,4) and β-(1,3) linkages with similar efficiency occurs when a β-(1,4) linkage of the heterogeneous polysaccharide can be placed between the −2 and −1 subsites. Additional studies with oligosaccharides have revealed subsite-by-subsite binding energies and rate constants for hydrolysis of simple oligosaccharides (9Meng D.D. Liu X. Dong S. Wang Y.F. Ma X.Q. Zhou H. Wang X. Yao L.S. Feng Y. Li F.L. Structural insights into the substrate specificity of a glycoside hydrolase family 5 lichenase from Caldicellulosiruptor sp. F32.Biochem. J. 2017; 474 (28838949): 3373-338910.1042/BCJ20170328Crossref PubMed Scopus (11) Google Scholar, 10Attia M.A. Nelson C.E. Offen W.A. Jain N. Davies G.J. Gardner J.G. Brumer H. In vitroin vivo characterization of three Cellvibrio japonicus glycoside hydrolase family 5 members reveals potent xyloglucan backbone-cleaving functions.Biotechnol. Biofuels. 2018; 11 (29467823): 4510.1186/s13068-018-1039-6Crossref PubMed Scopus (15) Google Scholar, 11McGregor N. Morar M. Fenger T.H. Stogios P. Lenfant N. Yin V. Xu X. Evdokimova E. Cui H. Henrissat B. Savchenko A. Brumer H. Structure-function analysis of a mixed-linkage β-glucanase/xyloglucanase from the key ruminal Bacteroidetes Prevotella bryantii B14.J. Biol. Chem. 2016; 291 (26507654): 1175-119710.1074/jbc.M115.691659Abstract Full Text Full Text PDF PubMed Scopus (29) Google Scholar, 12Tseng C.W. Ko T.P. Guo R.T. Huang J.W. Wang H.C. Huang C.H. Cheng Y.S. Wang A.H. Liu J.R. Substrate binding of a GH5 endoglucanase from the ruminal fungus Piromyces rhizinflata.Acta Crystallogr. Sect. F Struct. Biol. Cryst. Commun. 2011; 67 (22102024): 1189-119410.1107/S1744309111032428Crossref PubMed Scopus (21) Google Scholar, 16Dos Santos C.R. Cordeiro R.L. Wong D.W. Murakami M.T. Structural basis for xyloglucan specificity and α-d-Xylp(1 → 6)-d-Glcp recognition at the −1 subsite within the GH5 family.Biochemistry. 2015; 54 (25714929): 1930-194210.1021/acs.biochem.5b00011Crossref PubMed Scopus (13) Google Scholar, 17Gloster T.M. Ibatullin F.M. Macauley K. Eklöf J.M. Roberts S. Turkenburg J.P. Bjørnvad M.E. Jørgensen P.L. Danielsen S. Johansen K.S. Borchert T.V. Wilson K.S. Brumer H. Davies G.J. Characterization and three-dimensional structures of two distinct bacterial xyloglucanases from families GH5 and GH12.J. Biol. Chem. 2007; 282 (17376777): 19177-1918910.1074/jbc.M700224200Abstract Full Text Full Text PDF PubMed Scopus (86) Google Scholar, 18Venditto I. Najmudin S. Luis A.S. Ferreira L.M. Sakka K. Knox J.P. Gilbert H.J. Fontes C.M. Family 46 carbohydrate-binding modules contribute to the enzymatic hydrolysis of xyloglucan and β-1,3-1,4-glucans through distinct mechanisms.J. Biol. Chem. 2015; 290 (25713075): 10572-1058610.1074/jbc.M115.637827Abstract Full Text Full Text PDF PubMed Scopus (28) Google Scholar, 19Wu T.H. Huang C.H. Ko T.P. Lai H.L. Ma Y. Chen C.C. Cheng Y.S. Liu J.R. Guo R.T. Diverse substrate recognition mechanism revealed by Thermotoga maritima Cel5A structures in complex with cellotetraose, cellobiose and mannotriose.Biochim. Biophys. Acta. 2011; 1814 (21839861): 1832-184010.1016/j.bbapap.2011.07.020Crossref PubMed Scopus (28) Google Scholar, 20Yuan S.F. Wu T.H. Lee H.L. Hsieh H.Y. Lin W.L. Yang B. Chang C.K. Li Q. Gao J. Huang C.H. Ho M.C. Guo R.T. Liang P.H. Biochemical characterization and structural analysis of a bifunctional cellulase/xylanase from Clostridium thermocellum.J. Biol. Chem. 2015;
Referência(s)