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| | ==Multifunctional cellulase, xylanase, mannanase== | | ==Multifunctional cellulase, xylanase, mannanase== |
| - | <StructureSection load='4im4' size='340' side='right' caption='[[4im4]], [[Resolution|resolution]] 2.42Å' scene=''> | + | <StructureSection load='4im4' size='340' side='right'caption='[[4im4]], [[Resolution|resolution]] 2.42Å' scene=''> |
| | == Structural highlights == | | == Structural highlights == |
| - | <table><tr><td colspan='2'>[[4im4]] is a 6 chain structure with sequence from [http://en.wikipedia.org/wiki/"ruminiclostridium_thermocellum"_yutin_and_galperin_2013 "ruminiclostridium thermocellum" yutin and galperin 2013]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=4IM4 OCA]. For a <b>guided tour on the structure components</b> use [http://oca.weizmann.ac.il/oca-docs/fgij/fg.htm?mol=4IM4 FirstGlance]. <br> | + | <table><tr><td colspan='2'>[[4im4]] is a 6 chain structure with sequence from [https://en.wikipedia.org/wiki/Acetivibrio_thermocellus Acetivibrio thermocellus]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=4IM4 OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=4IM4 FirstGlance]. <br> |
| - | </td></tr><tr id='related'><td class="sblockLbl"><b>[[Related_structure|Related:]]</b></td><td class="sblockDat">[[4ilt|4ilt]], [[4ilv|4ilv]], [[4ily|4ily]], [[4im1|4im1]]</td></tr> | + | </td></tr><tr id='method'><td class="sblockLbl"><b>[[Empirical_models|Method:]]</b></td><td class="sblockDat" id="methodDat">X-ray diffraction, [[Resolution|Resolution]] 2.42Å</td></tr> |
| - | <tr id='gene'><td class="sblockLbl"><b>[[Gene|Gene:]]</b></td><td class="sblockDat">celE ([http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&srchmode=5&id=1515 "Ruminiclostridium thermocellum" Yutin and Galperin 2013])</td></tr>
| + | <tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[https://proteopedia.org/fgij/fg.htm?mol=4im4 FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=4im4 OCA], [https://pdbe.org/4im4 PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=4im4 RCSB], [https://www.ebi.ac.uk/pdbsum/4im4 PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=4im4 ProSAT]</span></td></tr> |
| - | <tr id='activity'><td class="sblockLbl"><b>Activity:</b></td><td class="sblockDat"><span class='plainlinks'>[http://en.wikipedia.org/wiki/Cellulase Cellulase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=3.2.1.4 3.2.1.4] </span></td></tr>
| + | |
| - | <tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[http://oca.weizmann.ac.il/oca-docs/fgij/fg.htm?mol=4im4 FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=4im4 OCA], [http://pdbe.org/4im4 PDBe], [http://www.rcsb.org/pdb/explore.do?structureId=4im4 RCSB], [http://www.ebi.ac.uk/pdbsum/4im4 PDBsum], [http://prosat.h-its.org/prosat/prosatexe?pdbcode=4im4 ProSAT]</span></td></tr> | + | |
| | </table> | | </table> |
| | + | == Function == |
| | + | [https://www.uniprot.org/uniprot/CELE_ACET2 CELE_ACET2] Multifunctional enzyme involved in the degradation of plant cell wall polysaccharides. Displays endoglucanase activity against carboxymethyl cellulose (CMC) and barley beta-glucan (PubMed:3066698, PubMed:1991028). Also catalyzes the deacetylation of acetylated birchwood xylan and glucomannan, with a preference for the latter, and of the synthetic substrate 4-nitrophenyl acetate (4-NPAc) (PubMed:19338387).<ref>PMID:19338387</ref> <ref>PMID:1991028</ref> <ref>PMID:3066698</ref> |
| | + | <div style="background-color:#fffaf0;"> |
| | + | == Publication Abstract from PubMed == |
| | + | 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 beta-(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 beta-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. |
| | + | |
| | + | A structural and kinetic survey of GH5_4 endoglucanases reveals determinants of broad substrate specificity and opportunities for biomass hydrolysis.,Glasgow EM, Kemna EI, Bingman CA, Ing N, Deng K, Bianchetti CM, Takasuka TE, Northen TR, Fox BG J Biol Chem. 2020 Dec 18;295(51):17752-17769. doi: 10.1074/jbc.RA120.015328. PMID:33454012<ref>PMID:33454012</ref> |
| | + | |
| | + | From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.<br> |
| | + | </div> |
| | + | <div class="pdbe-citations 4im4" style="background-color:#fffaf0;"></div> |
| | + | |
| | + | ==See Also== |
| | + | *[[Glucanase 3D structures|Glucanase 3D structures]] |
| | + | == References == |
| | + | <references/> |
| | __TOC__ | | __TOC__ |
| | </StructureSection> | | </StructureSection> |
| - | [[Category: Ruminiclostridium thermocellum yutin and galperin 2013]] | + | [[Category: Acetivibrio thermocellus]] |
| - | [[Category: Cellulase]] | + | [[Category: Large Structures]] |
| - | [[Category: Bianchetti, C M]] | + | [[Category: Bianchetti CM]] |
| - | [[Category: Fox, B G]] | + | [[Category: Fox BG]] |
| - | [[Category: Takasuka, T E]] | + | [[Category: Takasuka TE]] |
| - | [[Category: 4-beta-glucanase]]
| + | |
| - | [[Category: Biomass degradation]]
| + | |
| - | [[Category: Endo-1]]
| + | |
| - | [[Category: Hydrolase]]
| + | |
| - | [[Category: Mannanase]]
| + | |
| - | [[Category: Multifunction]]
| + | |
| - | [[Category: Xylanase]]
| + | |
| Structural highlights
Function
CELE_ACET2 Multifunctional enzyme involved in the degradation of plant cell wall polysaccharides. Displays endoglucanase activity against carboxymethyl cellulose (CMC) and barley beta-glucan (PubMed:3066698, PubMed:1991028). Also catalyzes the deacetylation of acetylated birchwood xylan and glucomannan, with a preference for the latter, and of the synthetic substrate 4-nitrophenyl acetate (4-NPAc) (PubMed:19338387).[1] [2] [3]
Publication Abstract from PubMed
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 beta-(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 beta-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.
A structural and kinetic survey of GH5_4 endoglucanases reveals determinants of broad substrate specificity and opportunities for biomass hydrolysis.,Glasgow EM, Kemna EI, Bingman CA, Ing N, Deng K, Bianchetti CM, Takasuka TE, Northen TR, Fox BG J Biol Chem. 2020 Dec 18;295(51):17752-17769. doi: 10.1074/jbc.RA120.015328. PMID:33454012[4]
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.
See Also
References
- ↑ Montanier C, Money VA, Pires VM, Flint JE, Pinheiro BA, Goyal A, Prates JA, Izumi A, Stalbrand H, Morland C, Cartmell A, Kolenova K, Topakas E, Dodson EJ, Bolam DN, Davies GJ, Fontes CM, Gilbert HJ. The active site of a carbohydrate esterase displays divergent catalytic and noncatalytic binding functions. PLoS Biol. 2009 Mar 31;7(3):e71. PMID:19338387 doi:10.1371/journal.pbio.1000071
- ↑ Durrant AJ, Hall J, Hazlewood GP, Gilbert HJ. The non-catalytic C-terminal region of endoglucanase E from Clostridium thermocellum contains a cellulose-binding domain. Biochem J. 1991 Jan 15;273(Pt 2):289-93. doi: 10.1042/bj2730289. PMID:1991028 doi:http://dx.doi.org/10.1042/bj2730289
- ↑ Hall J, Hazlewood GP, Barker PJ, Gilbert HJ. Conserved reiterated domains in Clostridium thermocellum endoglucanases are not essential for catalytic activity. Gene. 1988 Sep 15;69(1):29-38. PMID:3066698
- ↑ Glasgow EM, Kemna EI, Bingman CA, Ing N, Deng K, Bianchetti CM, Takasuka TE, Northen TR, Fox BG. A structural and kinetic survey of GH5_4 endoglucanases reveals determinants of broad substrate specificity and opportunities for biomass hydrolysis. J Biol Chem. 2020 Dec 18;295(51):17752-17769. doi: 10.1074/jbc.RA120.015328. PMID:33454012 doi:http://dx.doi.org/10.1074/jbc.RA120.015328
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