Unusual active site location and catalytic apparatus in a glycoside hydrolase family

The human gut microbiota use complex carbohydrates as major nutrients. The requirement for an efficient glycan degrading systems exerts a major selection pressure on this microbial community. Thus, we propose that these bacteria represent a substantial resource for discovering novel carbohydrate act...
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Autor*in:	Munoz-Munoz, Jose [verfasserIn] Cartmell, Alan Terrapon, Nicolas Henrissat, Bernard Gilbert, Harry J

Format:	Artikel
Sprache:	Englisch

Erschienen:	2017

Schlagwörter:	Glycosides Enzymes Hydrolases Microbiota (Symbiotic organisms) Research Hypothesis Usage Reaction products Bacteria Growth hormone Amino acids Residues a-L-Rhamnosidase Nutrients Site location Polysaccharides Mutagenesis Glycoside hydrolase Digestive tract Imidazole Histidine Hydrolase Crystal structure Carboxylic acids Microorganisms Carbohydrates L-Rhamnosidase Rhamnose

Übergeordnetes Werk:	Enthalten in: Proceedings of the National Academy of Sciences of the United States of America - Washington, DC : NAS, 1877, 114(2017), 19, Seite 4936
Übergeordnetes Werk:	volume:114 ; year:2017 ; number:19 ; pages:4936

Links:	Volltext Link aufrufen

DOI / URN:	10.1073/pnas.1701130114

Katalog-ID:	OLC1995553522

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520			\|a The human gut microbiota use complex carbohydrates as major nutrients. The requirement for an efficient glycan degrading systems exerts a major selection pressure on this microbial community. Thus, we propose that these bacteria represent a substantial resource for discovering novel carbohydrate active enzymes. To test this hypothesis, we focused on enzymes that hydrolyze rhamnosidic bonds, as cleavage of these linkages is chemically challenging and there is a paucity of information on L-rhamnosidases. Here we screened the activity of enzymes derived from the human gut microbiota bacterium Bacteroides thetaiotaomicron, which are up-regulated in response to rhamnose-containing glycans. We identified an a-L-rhamnosidase, BT3686, which is the founding member of a glycoside hydrolase (GH) family, GH145. In contrast to other rhamnosidases, BT3686 cleaved L-Rha-a1,4-D-GlcA linkages through a retaining double-displacement mechanism. The crystal structure of BT3686 showed that the enzyme displayed a type A seven-bladed β-propeller fold. Mutagenesis and crystallographic studies, including the structure of BT3686 in complex with the reaction product GlcA, revealed a location for the active site among β-propeller enzymes cited on the posterior surface of the rhamnosidase. In contrast to the vast majority of GH, the catalytic apparatus of BT3686 does not comprise a pair of carboxylic acid residues but, uniquely, a single histidine functions as the only discernable catalytic amino acid. Intriguingly, the histidine, His48, is not invariant in GH145; however, when engineered into structural homologs lacking the imidazole residue, a-L-rhamnosidase activity was established. The potential contribution of His48 to the catalytic activity of BT3686 is discussed.
650		4	\|a Glycosides
650		4	\|a Enzymes
650		4	\|a Hydrolases
650		4	\|a Microbiota (Symbiotic organisms)
650		4	\|a Research
650		4	\|a Hypothesis
650		4	\|a Usage
650		4	\|a Reaction products
650		4	\|a Bacteria
650		4	\|a Growth hormone
650		4	\|a Amino acids
650		4	\|a Residues
650		4	\|a a-L-Rhamnosidase
650		4	\|a Nutrients
650		4	\|a Site location
650		4	\|a Polysaccharides
650		4	\|a Mutagenesis
650		4	\|a Glycoside hydrolase
650		4	\|a Digestive tract
650		4	\|a Imidazole
650		4	\|a Histidine
650		4	\|a Hydrolase
650		4	\|a Crystal structure
650		4	\|a Carboxylic acids
650		4	\|a Microorganisms
650		4	\|a Carbohydrates
650		4	\|a L-Rhamnosidase
650		4	\|a Rhamnose
700	1		\|a Cartmell, Alan \|4 oth
700	1		\|a Terrapon, Nicolas \|4 oth
700	1		\|a Henrissat, Bernard \|4 oth
700	1		\|a Gilbert, Harry J \|4 oth
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allfields	10.1073/pnas.1701130114 doi PQ20170901 (DE-627)OLC1995553522 (DE-599)GBVOLC1995553522 (PRQ)g1160-ae592da50ac61fbc633e23c0802f6310cfa61710a1150a27107a5120b0f797150 (KEY)0583363920170000114001904936unusualactivesitelocationandcatalyticapparatusinag DE-627 ger DE-627 rakwb eng 500 DE-101 570 AVZ LING fid BIODIV fid Munoz-Munoz, Jose verfasserin aut Unusual active site location and catalytic apparatus in a glycoside hydrolase family 2017 Text txt rdacontent ohne Hilfsmittel zu benutzen n rdamedia Band nc rdacarrier The human gut microbiota use complex carbohydrates as major nutrients. The requirement for an efficient glycan degrading systems exerts a major selection pressure on this microbial community. Thus, we propose that these bacteria represent a substantial resource for discovering novel carbohydrate active enzymes. To test this hypothesis, we focused on enzymes that hydrolyze rhamnosidic bonds, as cleavage of these linkages is chemically challenging and there is a paucity of information on L-rhamnosidases. Here we screened the activity of enzymes derived from the human gut microbiota bacterium Bacteroides thetaiotaomicron, which are up-regulated in response to rhamnose-containing glycans. We identified an a-L-rhamnosidase, BT3686, which is the founding member of a glycoside hydrolase (GH) family, GH145. In contrast to other rhamnosidases, BT3686 cleaved L-Rha-a1,4-D-GlcA linkages through a retaining double-displacement mechanism. The crystal structure of BT3686 showed that the enzyme displayed a type A seven-bladed β-propeller fold. Mutagenesis and crystallographic studies, including the structure of BT3686 in complex with the reaction product GlcA, revealed a location for the active site among β-propeller enzymes cited on the posterior surface of the rhamnosidase. In contrast to the vast majority of GH, the catalytic apparatus of BT3686 does not comprise a pair of carboxylic acid residues but, uniquely, a single histidine functions as the only discernable catalytic amino acid. Intriguingly, the histidine, His48, is not invariant in GH145; however, when engineered into structural homologs lacking the imidazole residue, a-L-rhamnosidase activity was established. The potential contribution of His48 to the catalytic activity of BT3686 is discussed. Glycosides Enzymes Hydrolases Microbiota (Symbiotic organisms) Research Hypothesis Usage Reaction products Bacteria Growth hormone Amino acids Residues a-L-Rhamnosidase Nutrients Site location Polysaccharides Mutagenesis Glycoside hydrolase Digestive tract Imidazole Histidine Hydrolase Crystal structure Carboxylic acids Microorganisms Carbohydrates L-Rhamnosidase Rhamnose Cartmell, Alan oth Terrapon, Nicolas oth Henrissat, Bernard oth Gilbert, Harry J oth Enthalten in Proceedings of the National Academy of Sciences of the United States of America Washington, DC : NAS, 1877 114(2017), 19, Seite 4936 (DE-627)129505269 (DE-600)209104-5 (DE-576)014909189 0027-8424 nnns volume:114 year:2017 number:19 pages:4936 http://dx.doi.org/10.1073/pnas.1701130114 Volltext https://search.proquest.com/docview/1903952690 GBV_USEFLAG_A SYSFLAG_A GBV_OLC FID-LING FID-BIODIV SSG-OLC-PHY SSG-OLC-CHE SSG-OLC-MAT SSG-OLC-FOR SSG-OLC-PHA SSG-OLC-DE-84 SSG-OPC-MAT SSG-OPC-FOR GBV_ILN_40 GBV_ILN_59 AR 114 2017 19 4936
spelling	10.1073/pnas.1701130114 doi PQ20170901 (DE-627)OLC1995553522 (DE-599)GBVOLC1995553522 (PRQ)g1160-ae592da50ac61fbc633e23c0802f6310cfa61710a1150a27107a5120b0f797150 (KEY)0583363920170000114001904936unusualactivesitelocationandcatalyticapparatusinag DE-627 ger DE-627 rakwb eng 500 DE-101 570 AVZ LING fid BIODIV fid Munoz-Munoz, Jose verfasserin aut Unusual active site location and catalytic apparatus in a glycoside hydrolase family 2017 Text txt rdacontent ohne Hilfsmittel zu benutzen n rdamedia Band nc rdacarrier The human gut microbiota use complex carbohydrates as major nutrients. The requirement for an efficient glycan degrading systems exerts a major selection pressure on this microbial community. Thus, we propose that these bacteria represent a substantial resource for discovering novel carbohydrate active enzymes. To test this hypothesis, we focused on enzymes that hydrolyze rhamnosidic bonds, as cleavage of these linkages is chemically challenging and there is a paucity of information on L-rhamnosidases. Here we screened the activity of enzymes derived from the human gut microbiota bacterium Bacteroides thetaiotaomicron, which are up-regulated in response to rhamnose-containing glycans. We identified an a-L-rhamnosidase, BT3686, which is the founding member of a glycoside hydrolase (GH) family, GH145. In contrast to other rhamnosidases, BT3686 cleaved L-Rha-a1,4-D-GlcA linkages through a retaining double-displacement mechanism. The crystal structure of BT3686 showed that the enzyme displayed a type A seven-bladed β-propeller fold. Mutagenesis and crystallographic studies, including the structure of BT3686 in complex with the reaction product GlcA, revealed a location for the active site among β-propeller enzymes cited on the posterior surface of the rhamnosidase. In contrast to the vast majority of GH, the catalytic apparatus of BT3686 does not comprise a pair of carboxylic acid residues but, uniquely, a single histidine functions as the only discernable catalytic amino acid. Intriguingly, the histidine, His48, is not invariant in GH145; however, when engineered into structural homologs lacking the imidazole residue, a-L-rhamnosidase activity was established. The potential contribution of His48 to the catalytic activity of BT3686 is discussed. Glycosides Enzymes Hydrolases Microbiota (Symbiotic organisms) Research Hypothesis Usage Reaction products Bacteria Growth hormone Amino acids Residues a-L-Rhamnosidase Nutrients Site location Polysaccharides Mutagenesis Glycoside hydrolase Digestive tract Imidazole Histidine Hydrolase Crystal structure Carboxylic acids Microorganisms Carbohydrates L-Rhamnosidase Rhamnose Cartmell, Alan oth Terrapon, Nicolas oth Henrissat, Bernard oth Gilbert, Harry J oth Enthalten in Proceedings of the National Academy of Sciences of the United States of America Washington, DC : NAS, 1877 114(2017), 19, Seite 4936 (DE-627)129505269 (DE-600)209104-5 (DE-576)014909189 0027-8424 nnns volume:114 year:2017 number:19 pages:4936 http://dx.doi.org/10.1073/pnas.1701130114 Volltext https://search.proquest.com/docview/1903952690 GBV_USEFLAG_A SYSFLAG_A GBV_OLC FID-LING FID-BIODIV SSG-OLC-PHY SSG-OLC-CHE SSG-OLC-MAT SSG-OLC-FOR SSG-OLC-PHA SSG-OLC-DE-84 SSG-OPC-MAT SSG-OPC-FOR GBV_ILN_40 GBV_ILN_59 AR 114 2017 19 4936
allfields_unstemmed	10.1073/pnas.1701130114 doi PQ20170901 (DE-627)OLC1995553522 (DE-599)GBVOLC1995553522 (PRQ)g1160-ae592da50ac61fbc633e23c0802f6310cfa61710a1150a27107a5120b0f797150 (KEY)0583363920170000114001904936unusualactivesitelocationandcatalyticapparatusinag DE-627 ger DE-627 rakwb eng 500 DE-101 570 AVZ LING fid BIODIV fid Munoz-Munoz, Jose verfasserin aut Unusual active site location and catalytic apparatus in a glycoside hydrolase family 2017 Text txt rdacontent ohne Hilfsmittel zu benutzen n rdamedia Band nc rdacarrier The human gut microbiota use complex carbohydrates as major nutrients. The requirement for an efficient glycan degrading systems exerts a major selection pressure on this microbial community. Thus, we propose that these bacteria represent a substantial resource for discovering novel carbohydrate active enzymes. To test this hypothesis, we focused on enzymes that hydrolyze rhamnosidic bonds, as cleavage of these linkages is chemically challenging and there is a paucity of information on L-rhamnosidases. Here we screened the activity of enzymes derived from the human gut microbiota bacterium Bacteroides thetaiotaomicron, which are up-regulated in response to rhamnose-containing glycans. We identified an a-L-rhamnosidase, BT3686, which is the founding member of a glycoside hydrolase (GH) family, GH145. In contrast to other rhamnosidases, BT3686 cleaved L-Rha-a1,4-D-GlcA linkages through a retaining double-displacement mechanism. The crystal structure of BT3686 showed that the enzyme displayed a type A seven-bladed β-propeller fold. Mutagenesis and crystallographic studies, including the structure of BT3686 in complex with the reaction product GlcA, revealed a location for the active site among β-propeller enzymes cited on the posterior surface of the rhamnosidase. In contrast to the vast majority of GH, the catalytic apparatus of BT3686 does not comprise a pair of carboxylic acid residues but, uniquely, a single histidine functions as the only discernable catalytic amino acid. Intriguingly, the histidine, His48, is not invariant in GH145; however, when engineered into structural homologs lacking the imidazole residue, a-L-rhamnosidase activity was established. The potential contribution of His48 to the catalytic activity of BT3686 is discussed. Glycosides Enzymes Hydrolases Microbiota (Symbiotic organisms) Research Hypothesis Usage Reaction products Bacteria Growth hormone Amino acids Residues a-L-Rhamnosidase Nutrients Site location Polysaccharides Mutagenesis Glycoside hydrolase Digestive tract Imidazole Histidine Hydrolase Crystal structure Carboxylic acids Microorganisms Carbohydrates L-Rhamnosidase Rhamnose Cartmell, Alan oth Terrapon, Nicolas oth Henrissat, Bernard oth Gilbert, Harry J oth Enthalten in Proceedings of the National Academy of Sciences of the United States of America Washington, DC : NAS, 1877 114(2017), 19, Seite 4936 (DE-627)129505269 (DE-600)209104-5 (DE-576)014909189 0027-8424 nnns volume:114 year:2017 number:19 pages:4936 http://dx.doi.org/10.1073/pnas.1701130114 Volltext https://search.proquest.com/docview/1903952690 GBV_USEFLAG_A SYSFLAG_A GBV_OLC FID-LING FID-BIODIV SSG-OLC-PHY SSG-OLC-CHE SSG-OLC-MAT SSG-OLC-FOR SSG-OLC-PHA SSG-OLC-DE-84 SSG-OPC-MAT SSG-OPC-FOR GBV_ILN_40 GBV_ILN_59 AR 114 2017 19 4936
allfieldsGer	10.1073/pnas.1701130114 doi PQ20170901 (DE-627)OLC1995553522 (DE-599)GBVOLC1995553522 (PRQ)g1160-ae592da50ac61fbc633e23c0802f6310cfa61710a1150a27107a5120b0f797150 (KEY)0583363920170000114001904936unusualactivesitelocationandcatalyticapparatusinag DE-627 ger DE-627 rakwb eng 500 DE-101 570 AVZ LING fid BIODIV fid Munoz-Munoz, Jose verfasserin aut Unusual active site location and catalytic apparatus in a glycoside hydrolase family 2017 Text txt rdacontent ohne Hilfsmittel zu benutzen n rdamedia Band nc rdacarrier The human gut microbiota use complex carbohydrates as major nutrients. The requirement for an efficient glycan degrading systems exerts a major selection pressure on this microbial community. Thus, we propose that these bacteria represent a substantial resource for discovering novel carbohydrate active enzymes. To test this hypothesis, we focused on enzymes that hydrolyze rhamnosidic bonds, as cleavage of these linkages is chemically challenging and there is a paucity of information on L-rhamnosidases. Here we screened the activity of enzymes derived from the human gut microbiota bacterium Bacteroides thetaiotaomicron, which are up-regulated in response to rhamnose-containing glycans. We identified an a-L-rhamnosidase, BT3686, which is the founding member of a glycoside hydrolase (GH) family, GH145. In contrast to other rhamnosidases, BT3686 cleaved L-Rha-a1,4-D-GlcA linkages through a retaining double-displacement mechanism. The crystal structure of BT3686 showed that the enzyme displayed a type A seven-bladed β-propeller fold. Mutagenesis and crystallographic studies, including the structure of BT3686 in complex with the reaction product GlcA, revealed a location for the active site among β-propeller enzymes cited on the posterior surface of the rhamnosidase. In contrast to the vast majority of GH, the catalytic apparatus of BT3686 does not comprise a pair of carboxylic acid residues but, uniquely, a single histidine functions as the only discernable catalytic amino acid. Intriguingly, the histidine, His48, is not invariant in GH145; however, when engineered into structural homologs lacking the imidazole residue, a-L-rhamnosidase activity was established. The potential contribution of His48 to the catalytic activity of BT3686 is discussed. Glycosides Enzymes Hydrolases Microbiota (Symbiotic organisms) Research Hypothesis Usage Reaction products Bacteria Growth hormone Amino acids Residues a-L-Rhamnosidase Nutrients Site location Polysaccharides Mutagenesis Glycoside hydrolase Digestive tract Imidazole Histidine Hydrolase Crystal structure Carboxylic acids Microorganisms Carbohydrates L-Rhamnosidase Rhamnose Cartmell, Alan oth Terrapon, Nicolas oth Henrissat, Bernard oth Gilbert, Harry J oth Enthalten in Proceedings of the National Academy of Sciences of the United States of America Washington, DC : NAS, 1877 114(2017), 19, Seite 4936 (DE-627)129505269 (DE-600)209104-5 (DE-576)014909189 0027-8424 nnns volume:114 year:2017 number:19 pages:4936 http://dx.doi.org/10.1073/pnas.1701130114 Volltext https://search.proquest.com/docview/1903952690 GBV_USEFLAG_A SYSFLAG_A GBV_OLC FID-LING FID-BIODIV SSG-OLC-PHY SSG-OLC-CHE SSG-OLC-MAT SSG-OLC-FOR SSG-OLC-PHA SSG-OLC-DE-84 SSG-OPC-MAT SSG-OPC-FOR GBV_ILN_40 GBV_ILN_59 AR 114 2017 19 4936
allfieldsSound	10.1073/pnas.1701130114 doi PQ20170901 (DE-627)OLC1995553522 (DE-599)GBVOLC1995553522 (PRQ)g1160-ae592da50ac61fbc633e23c0802f6310cfa61710a1150a27107a5120b0f797150 (KEY)0583363920170000114001904936unusualactivesitelocationandcatalyticapparatusinag DE-627 ger DE-627 rakwb eng 500 DE-101 570 AVZ LING fid BIODIV fid Munoz-Munoz, Jose verfasserin aut Unusual active site location and catalytic apparatus in a glycoside hydrolase family 2017 Text txt rdacontent ohne Hilfsmittel zu benutzen n rdamedia Band nc rdacarrier The human gut microbiota use complex carbohydrates as major nutrients. The requirement for an efficient glycan degrading systems exerts a major selection pressure on this microbial community. Thus, we propose that these bacteria represent a substantial resource for discovering novel carbohydrate active enzymes. To test this hypothesis, we focused on enzymes that hydrolyze rhamnosidic bonds, as cleavage of these linkages is chemically challenging and there is a paucity of information on L-rhamnosidases. Here we screened the activity of enzymes derived from the human gut microbiota bacterium Bacteroides thetaiotaomicron, which are up-regulated in response to rhamnose-containing glycans. We identified an a-L-rhamnosidase, BT3686, which is the founding member of a glycoside hydrolase (GH) family, GH145. In contrast to other rhamnosidases, BT3686 cleaved L-Rha-a1,4-D-GlcA linkages through a retaining double-displacement mechanism. The crystal structure of BT3686 showed that the enzyme displayed a type A seven-bladed β-propeller fold. Mutagenesis and crystallographic studies, including the structure of BT3686 in complex with the reaction product GlcA, revealed a location for the active site among β-propeller enzymes cited on the posterior surface of the rhamnosidase. In contrast to the vast majority of GH, the catalytic apparatus of BT3686 does not comprise a pair of carboxylic acid residues but, uniquely, a single histidine functions as the only discernable catalytic amino acid. Intriguingly, the histidine, His48, is not invariant in GH145; however, when engineered into structural homologs lacking the imidazole residue, a-L-rhamnosidase activity was established. The potential contribution of His48 to the catalytic activity of BT3686 is discussed. Glycosides Enzymes Hydrolases Microbiota (Symbiotic organisms) Research Hypothesis Usage Reaction products Bacteria Growth hormone Amino acids Residues a-L-Rhamnosidase Nutrients Site location Polysaccharides Mutagenesis Glycoside hydrolase Digestive tract Imidazole Histidine Hydrolase Crystal structure Carboxylic acids Microorganisms Carbohydrates L-Rhamnosidase Rhamnose Cartmell, Alan oth Terrapon, Nicolas oth Henrissat, Bernard oth Gilbert, Harry J oth Enthalten in Proceedings of the National Academy of Sciences of the United States of America Washington, DC : NAS, 1877 114(2017), 19, Seite 4936 (DE-627)129505269 (DE-600)209104-5 (DE-576)014909189 0027-8424 nnns volume:114 year:2017 number:19 pages:4936 http://dx.doi.org/10.1073/pnas.1701130114 Volltext https://search.proquest.com/docview/1903952690 GBV_USEFLAG_A SYSFLAG_A GBV_OLC FID-LING FID-BIODIV SSG-OLC-PHY SSG-OLC-CHE SSG-OLC-MAT SSG-OLC-FOR SSG-OLC-PHA SSG-OLC-DE-84 SSG-OPC-MAT SSG-OPC-FOR GBV_ILN_40 GBV_ILN_59 AR 114 2017 19 4936
language	English
source	Enthalten in Proceedings of the National Academy of Sciences of the United States of America 114(2017), 19, Seite 4936 volume:114 year:2017 number:19 pages:4936
sourceStr	Enthalten in Proceedings of the National Academy of Sciences of the United States of America 114(2017), 19, Seite 4936 volume:114 year:2017 number:19 pages:4936
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topic_facet	Glycosides Enzymes Hydrolases Microbiota (Symbiotic organisms) Research Hypothesis Usage Reaction products Bacteria Growth hormone Amino acids Residues a-L-Rhamnosidase Nutrients Site location Polysaccharides Mutagenesis Glycoside hydrolase Digestive tract Imidazole Histidine Hydrolase Crystal structure Carboxylic acids Microorganisms Carbohydrates L-Rhamnosidase Rhamnose
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author	Munoz-Munoz, Jose
spellingShingle	Munoz-Munoz, Jose ddc 500 ddc 570 fid LING fid BIODIV misc Glycosides misc Enzymes misc Hydrolases misc Microbiota (Symbiotic organisms) misc Research misc Hypothesis misc Usage misc Reaction products misc Bacteria misc Growth hormone misc Amino acids misc Residues misc a-L-Rhamnosidase misc Nutrients misc Site location misc Polysaccharides misc Mutagenesis misc Glycoside hydrolase misc Digestive tract misc Imidazole misc Histidine misc Hydrolase misc Crystal structure misc Carboxylic acids misc Microorganisms misc Carbohydrates misc L-Rhamnosidase misc Rhamnose Unusual active site location and catalytic apparatus in a glycoside hydrolase family
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topic_title	500 DE-101 570 AVZ LING fid BIODIV fid Unusual active site location and catalytic apparatus in a glycoside hydrolase family Glycosides Enzymes Hydrolases Microbiota (Symbiotic organisms) Research Hypothesis Usage Reaction products Bacteria Growth hormone Amino acids Residues a-L-Rhamnosidase Nutrients Site location Polysaccharides Mutagenesis Glycoside hydrolase Digestive tract Imidazole Histidine Hydrolase Crystal structure Carboxylic acids Microorganisms Carbohydrates L-Rhamnosidase Rhamnose
topic	ddc 500 ddc 570 fid LING fid BIODIV misc Glycosides misc Enzymes misc Hydrolases misc Microbiota (Symbiotic organisms) misc Research misc Hypothesis misc Usage misc Reaction products misc Bacteria misc Growth hormone misc Amino acids misc Residues misc a-L-Rhamnosidase misc Nutrients misc Site location misc Polysaccharides misc Mutagenesis misc Glycoside hydrolase misc Digestive tract misc Imidazole misc Histidine misc Hydrolase misc Crystal structure misc Carboxylic acids misc Microorganisms misc Carbohydrates misc L-Rhamnosidase misc Rhamnose
topic_unstemmed	ddc 500 ddc 570 fid LING fid BIODIV misc Glycosides misc Enzymes misc Hydrolases misc Microbiota (Symbiotic organisms) misc Research misc Hypothesis misc Usage misc Reaction products misc Bacteria misc Growth hormone misc Amino acids misc Residues misc a-L-Rhamnosidase misc Nutrients misc Site location misc Polysaccharides misc Mutagenesis misc Glycoside hydrolase misc Digestive tract misc Imidazole misc Histidine misc Hydrolase misc Crystal structure misc Carboxylic acids misc Microorganisms misc Carbohydrates misc L-Rhamnosidase misc Rhamnose
topic_browse	ddc 500 ddc 570 fid LING fid BIODIV misc Glycosides misc Enzymes misc Hydrolases misc Microbiota (Symbiotic organisms) misc Research misc Hypothesis misc Usage misc Reaction products misc Bacteria misc Growth hormone misc Amino acids misc Residues misc a-L-Rhamnosidase misc Nutrients misc Site location misc Polysaccharides misc Mutagenesis misc Glycoside hydrolase misc Digestive tract misc Imidazole misc Histidine misc Hydrolase misc Crystal structure misc Carboxylic acids misc Microorganisms misc Carbohydrates misc L-Rhamnosidase misc Rhamnose
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title	Unusual active site location and catalytic apparatus in a glycoside hydrolase family
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title_full	Unusual active site location and catalytic apparatus in a glycoside hydrolase family
author_sort	Munoz-Munoz, Jose
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title_sort	unusual active site location and catalytic apparatus in a glycoside hydrolase family
title_auth	Unusual active site location and catalytic apparatus in a glycoside hydrolase family
abstract	The human gut microbiota use complex carbohydrates as major nutrients. The requirement for an efficient glycan degrading systems exerts a major selection pressure on this microbial community. Thus, we propose that these bacteria represent a substantial resource for discovering novel carbohydrate active enzymes. To test this hypothesis, we focused on enzymes that hydrolyze rhamnosidic bonds, as cleavage of these linkages is chemically challenging and there is a paucity of information on L-rhamnosidases. Here we screened the activity of enzymes derived from the human gut microbiota bacterium Bacteroides thetaiotaomicron, which are up-regulated in response to rhamnose-containing glycans. We identified an a-L-rhamnosidase, BT3686, which is the founding member of a glycoside hydrolase (GH) family, GH145. In contrast to other rhamnosidases, BT3686 cleaved L-Rha-a1,4-D-GlcA linkages through a retaining double-displacement mechanism. The crystal structure of BT3686 showed that the enzyme displayed a type A seven-bladed β-propeller fold. Mutagenesis and crystallographic studies, including the structure of BT3686 in complex with the reaction product GlcA, revealed a location for the active site among β-propeller enzymes cited on the posterior surface of the rhamnosidase. In contrast to the vast majority of GH, the catalytic apparatus of BT3686 does not comprise a pair of carboxylic acid residues but, uniquely, a single histidine functions as the only discernable catalytic amino acid. Intriguingly, the histidine, His48, is not invariant in GH145; however, when engineered into structural homologs lacking the imidazole residue, a-L-rhamnosidase activity was established. The potential contribution of His48 to the catalytic activity of BT3686 is discussed.
abstractGer	The human gut microbiota use complex carbohydrates as major nutrients. The requirement for an efficient glycan degrading systems exerts a major selection pressure on this microbial community. Thus, we propose that these bacteria represent a substantial resource for discovering novel carbohydrate active enzymes. To test this hypothesis, we focused on enzymes that hydrolyze rhamnosidic bonds, as cleavage of these linkages is chemically challenging and there is a paucity of information on L-rhamnosidases. Here we screened the activity of enzymes derived from the human gut microbiota bacterium Bacteroides thetaiotaomicron, which are up-regulated in response to rhamnose-containing glycans. We identified an a-L-rhamnosidase, BT3686, which is the founding member of a glycoside hydrolase (GH) family, GH145. In contrast to other rhamnosidases, BT3686 cleaved L-Rha-a1,4-D-GlcA linkages through a retaining double-displacement mechanism. The crystal structure of BT3686 showed that the enzyme displayed a type A seven-bladed β-propeller fold. Mutagenesis and crystallographic studies, including the structure of BT3686 in complex with the reaction product GlcA, revealed a location for the active site among β-propeller enzymes cited on the posterior surface of the rhamnosidase. In contrast to the vast majority of GH, the catalytic apparatus of BT3686 does not comprise a pair of carboxylic acid residues but, uniquely, a single histidine functions as the only discernable catalytic amino acid. Intriguingly, the histidine, His48, is not invariant in GH145; however, when engineered into structural homologs lacking the imidazole residue, a-L-rhamnosidase activity was established. The potential contribution of His48 to the catalytic activity of BT3686 is discussed.
abstract_unstemmed	The human gut microbiota use complex carbohydrates as major nutrients. The requirement for an efficient glycan degrading systems exerts a major selection pressure on this microbial community. Thus, we propose that these bacteria represent a substantial resource for discovering novel carbohydrate active enzymes. To test this hypothesis, we focused on enzymes that hydrolyze rhamnosidic bonds, as cleavage of these linkages is chemically challenging and there is a paucity of information on L-rhamnosidases. Here we screened the activity of enzymes derived from the human gut microbiota bacterium Bacteroides thetaiotaomicron, which are up-regulated in response to rhamnose-containing glycans. We identified an a-L-rhamnosidase, BT3686, which is the founding member of a glycoside hydrolase (GH) family, GH145. In contrast to other rhamnosidases, BT3686 cleaved L-Rha-a1,4-D-GlcA linkages through a retaining double-displacement mechanism. The crystal structure of BT3686 showed that the enzyme displayed a type A seven-bladed β-propeller fold. Mutagenesis and crystallographic studies, including the structure of BT3686 in complex with the reaction product GlcA, revealed a location for the active site among β-propeller enzymes cited on the posterior surface of the rhamnosidase. In contrast to the vast majority of GH, the catalytic apparatus of BT3686 does not comprise a pair of carboxylic acid residues but, uniquely, a single histidine functions as the only discernable catalytic amino acid. Intriguingly, the histidine, His48, is not invariant in GH145; however, when engineered into structural homologs lacking the imidazole residue, a-L-rhamnosidase activity was established. The potential contribution of His48 to the catalytic activity of BT3686 is discussed.
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title_short	Unusual active site location and catalytic apparatus in a glycoside hydrolase family
url	http://dx.doi.org/10.1073/pnas.1701130114 https://search.proquest.com/docview/1903952690
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Mutagenesis and crystallographic studies, including the structure of BT3686 in complex with the reaction product GlcA, revealed a location for the active site among β-propeller enzymes cited on the posterior surface of the rhamnosidase. In contrast to the vast majority of GH, the catalytic apparatus of BT3686 does not comprise a pair of carboxylic acid residues but, uniquely, a single histidine functions as the only discernable catalytic amino acid. Intriguingly, the histidine, His48, is not invariant in GH145; however, when engineered into structural homologs lacking the imidazole residue, a-L-rhamnosidase activity was established. 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tag="650" ind1=" " ind2="4"><subfield code="a">Histidine</subfield></datafield><datafield tag="650" ind1=" " ind2="4"><subfield code="a">Hydrolase</subfield></datafield><datafield tag="650" ind1=" " ind2="4"><subfield code="a">Crystal structure</subfield></datafield><datafield tag="650" ind1=" " ind2="4"><subfield code="a">Carboxylic acids</subfield></datafield><datafield tag="650" ind1=" " ind2="4"><subfield code="a">Microorganisms</subfield></datafield><datafield tag="650" ind1=" " ind2="4"><subfield code="a">Carbohydrates</subfield></datafield><datafield tag="650" ind1=" " ind2="4"><subfield code="a">L-Rhamnosidase</subfield></datafield><datafield tag="650" ind1=" " ind2="4"><subfield code="a">Rhamnose</subfield></datafield><datafield tag="700" ind1="1" ind2=" "><subfield code="a">Cartmell, Alan</subfield><subfield code="4">oth</subfield></datafield><datafield tag="700" ind1="1" ind2=" "><subfield code="a">Terrapon, Nicolas</subfield><subfield code="4">oth</subfield></datafield><datafield tag="700" ind1="1" ind2=" "><subfield code="a">Henrissat, Bernard</subfield><subfield code="4">oth</subfield></datafield><datafield tag="700" ind1="1" ind2=" "><subfield code="a">Gilbert, Harry J</subfield><subfield code="4">oth</subfield></datafield><datafield tag="773" ind1="0" ind2="8"><subfield code="i">Enthalten in</subfield><subfield code="t">Proceedings of the National Academy of Sciences of the United States of America</subfield><subfield code="d">Washington, DC : NAS, 1877</subfield><subfield code="g">114(2017), 19, Seite 4936</subfield><subfield code="w">(DE-627)129505269</subfield><subfield code="w">(DE-600)209104-5</subfield><subfield code="w">(DE-576)014909189</subfield><subfield code="x">0027-8424</subfield><subfield code="7">nnns</subfield></datafield><datafield tag="773" ind1="1" ind2="8"><subfield code="g">volume:114</subfield><subfield code="g">year:2017</subfield><subfield code="g">number:19</subfield><subfield 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Unusual active site location and catalytic apparatus in a glycoside hydrolase family

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