Structure of the Humicola insolens cellobiohydrolase Cel6A D416A mutant in complex with a non-hydrolysable substrate analogue, methyl cellobiosyl-4-thio-beta-cellobioside, at 1.9 angstrom

A  Varrot; T P  Frandsen; H  Driguez; G J  Davies

doi:10.1107/S0907444902017006

Structure of the Humicola insolens cellobiohydrolase Cel6A D416A mutant in complex with a non-hydrolysable substrate analogue, methyl cellobiosyl-4-thio-beta-cellobioside, at 1.9 angstrom

A Varrot, T P Frandsen, H Driguez, G J Davies

Chemistry

Research output: Contribution to journal › Article › peer-review

Abstract

The enzymatic degradation of cellulose continues to be one of the most important enzyme-catalysed reactions. Glycoside hydrolases from family GH-6 hydrolyse cellulose with inversion of the configuration of the anomeric carbon. Whilst the catalytic proton donor has been clearly identified (Asp226 in Humicola insolens Cel6A), the identification and even the existence of a potential Bronsted base remains unclear. Equally controversial is the role of surface-loop flexibility. Here, the structure of the D416A mutant of the H. insolens cellobiohydrolase Cel6A in complex with a nonhydrolysable thiooligosaccharide methyl cellobiosyl-4-thio-beta-cellobioside at 1.9 Angstrom resolution is presented. Substrate distortion in the -1 subsite, to a S-2(0) skew-boat conformation, is observed, similar to that seen in the analogous Trichoderma reesei Cel6A structure [Zou et al. (1999), Structure, 7, 1035-1045], but the active-centre N-terminal loop of the H. insolens enzyme is found in a more open conformation than described for previous structures.

Original language	English
Pages (from-to)	2201-2204
Number of pages	4
Journal	Acta Crystallographica. Section D, Biological Crystallography
Volume	58
DOIs	https://doi.org/10.1107/S0907444902017006
Publication status	Published - Dec 2002

Keywords

GLYCOSYL-ENZYME INTERMEDIATE
TRICHODERMA-REESEI
ACTIVE-SITE
OLIGOSACCHARIDE BINDING
CRYSTAL-STRUCTURE
CATALYTIC DOMAIN
RING DISTORTION
ENDOGLUCANASE
SPECIFICITY
FAMILY-6

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Cite this

Varrot, A., Frandsen, T. P., Driguez, H., & Davies, G. J. (2002). Structure of the Humicola insolens cellobiohydrolase Cel6A D416A mutant in complex with a non-hydrolysable substrate analogue, methyl cellobiosyl-4-thio-beta-cellobioside, at 1.9 angstrom. Acta Crystallographica. Section D, Biological Crystallography, 58, 2201-2204. https://doi.org/10.1107/S0907444902017006

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title = "Structure of the Humicola insolens cellobiohydrolase Cel6A D416A mutant in complex with a non-hydrolysable substrate analogue, methyl cellobiosyl-4-thio-beta-cellobioside, at 1.9 angstrom",

abstract = "The enzymatic degradation of cellulose continues to be one of the most important enzyme-catalysed reactions. Glycoside hydrolases from family GH-6 hydrolyse cellulose with inversion of the configuration of the anomeric carbon. Whilst the catalytic proton donor has been clearly identified (Asp226 in Humicola insolens Cel6A), the identification and even the existence of a potential Bronsted base remains unclear. Equally controversial is the role of surface-loop flexibility. Here, the structure of the D416A mutant of the H. insolens cellobiohydrolase Cel6A in complex with a nonhydrolysable thiooligosaccharide methyl cellobiosyl-4-thio-beta-cellobioside at 1.9 Angstrom resolution is presented. Substrate distortion in the -1 subsite, to a S-2(0) skew-boat conformation, is observed, similar to that seen in the analogous Trichoderma reesei Cel6A structure [Zou et al. (1999), Structure, 7, 1035-1045], but the active-centre N-terminal loop of the H. insolens enzyme is found in a more open conformation than described for previous structures.",

keywords = "GLYCOSYL-ENZYME INTERMEDIATE, TRICHODERMA-REESEI, ACTIVE-SITE, OLIGOSACCHARIDE BINDING, CRYSTAL-STRUCTURE, CATALYTIC DOMAIN, RING DISTORTION, ENDOGLUCANASE, SPECIFICITY, FAMILY-6",

author = "A Varrot and Frandsen, {T P} and H Driguez and Davies, {G J}",

year = "2002",

month = dec,

doi = "10.1107/S0907444902017006",

language = "English",

volume = "58",

pages = "2201--2204",

journal = "Acta Crystallographica. Section D, Biological Crystallography",

issn = "0907-4449",

publisher = "International Union of Crystallography",

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Varrot, A, Frandsen, TP, Driguez, H & Davies, GJ 2002, 'Structure of the Humicola insolens cellobiohydrolase Cel6A D416A mutant in complex with a non-hydrolysable substrate analogue, methyl cellobiosyl-4-thio-beta-cellobioside, at 1.9 angstrom', Acta Crystallographica. Section D, Biological Crystallography, vol. 58, pp. 2201-2204. https://doi.org/10.1107/S0907444902017006

Structure of the Humicola insolens cellobiohydrolase Cel6A D416A mutant in complex with a non-hydrolysable substrate analogue, methyl cellobiosyl-4-thio-beta-cellobioside, at 1.9 angstrom. / Varrot, A ; Frandsen, T P ; Driguez, H et al.
In: Acta Crystallographica. Section D, Biological Crystallography, Vol. 58, 12.2002, p. 2201-2204.

Research output: Contribution to journal › Article › peer-review

TY - JOUR

T1 - Structure of the Humicola insolens cellobiohydrolase Cel6A D416A mutant in complex with a non-hydrolysable substrate analogue, methyl cellobiosyl-4-thio-beta-cellobioside, at 1.9 angstrom

AU - Varrot, A

AU - Frandsen, T P

AU - Driguez, H

AU - Davies, G J

PY - 2002/12

Y1 - 2002/12

N2 - The enzymatic degradation of cellulose continues to be one of the most important enzyme-catalysed reactions. Glycoside hydrolases from family GH-6 hydrolyse cellulose with inversion of the configuration of the anomeric carbon. Whilst the catalytic proton donor has been clearly identified (Asp226 in Humicola insolens Cel6A), the identification and even the existence of a potential Bronsted base remains unclear. Equally controversial is the role of surface-loop flexibility. Here, the structure of the D416A mutant of the H. insolens cellobiohydrolase Cel6A in complex with a nonhydrolysable thiooligosaccharide methyl cellobiosyl-4-thio-beta-cellobioside at 1.9 Angstrom resolution is presented. Substrate distortion in the -1 subsite, to a S-2(0) skew-boat conformation, is observed, similar to that seen in the analogous Trichoderma reesei Cel6A structure [Zou et al. (1999), Structure, 7, 1035-1045], but the active-centre N-terminal loop of the H. insolens enzyme is found in a more open conformation than described for previous structures.

AB - The enzymatic degradation of cellulose continues to be one of the most important enzyme-catalysed reactions. Glycoside hydrolases from family GH-6 hydrolyse cellulose with inversion of the configuration of the anomeric carbon. Whilst the catalytic proton donor has been clearly identified (Asp226 in Humicola insolens Cel6A), the identification and even the existence of a potential Bronsted base remains unclear. Equally controversial is the role of surface-loop flexibility. Here, the structure of the D416A mutant of the H. insolens cellobiohydrolase Cel6A in complex with a nonhydrolysable thiooligosaccharide methyl cellobiosyl-4-thio-beta-cellobioside at 1.9 Angstrom resolution is presented. Substrate distortion in the -1 subsite, to a S-2(0) skew-boat conformation, is observed, similar to that seen in the analogous Trichoderma reesei Cel6A structure [Zou et al. (1999), Structure, 7, 1035-1045], but the active-centre N-terminal loop of the H. insolens enzyme is found in a more open conformation than described for previous structures.

KW - GLYCOSYL-ENZYME INTERMEDIATE

KW - TRICHODERMA-REESEI

KW - ACTIVE-SITE

KW - OLIGOSACCHARIDE BINDING

KW - CRYSTAL-STRUCTURE

KW - CATALYTIC DOMAIN

KW - RING DISTORTION

KW - ENDOGLUCANASE

KW - SPECIFICITY

KW - FAMILY-6

U2 - 10.1107/S0907444902017006

DO - 10.1107/S0907444902017006

M3 - Article

SN - 0907-4449

VL - 58

SP - 2201

EP - 2204

JO - Acta Crystallographica. Section D, Biological Crystallography

JF - Acta Crystallographica. Section D, Biological Crystallography

ER -