TY - JOUR

T1 - Sequencing, biochemical characterization, crystal structure and molecular dynamics of cellobiohydrolase Cel7A from Geotrichum candidum 3C

AU - Borisova, Anna S.

AU - Eneyskaya, Elena V.

AU - Bobrov, Kirill S.

AU - Jana, Suvamay

AU - Logachev, Anton

AU - Polev, Dmitrii E.

AU - Lapidus, Alla L.

AU - Ibatullin, Farid M.

AU - Saleem, Umair

AU - Sandgren, Mats

AU - Payne, Christina M.

AU - Kulminskaya, Anna A.

AU - Stahlberg, Jerry

PY - 2015/12

Y1 - 2015/12

N2 - The ascomycete Geotrichum candidum is a versatile and efficient decay fungus that is involved, for example, in biodeterioration of compact discs; notably, the 3C strain was previously shown to degrade filter paper and cotton more efficiently than several industrial enzyme preparations. Glycoside hydrolase (GH) family 7 cellobiohydrolases (CBHs) are the primary constituents of industrial cellulase cocktails employed in biomass conversion, and feature tunnel-enclosed active sites that enable processive hydrolytic cleavage of cellulose chains. Understanding the structure-function relationships defining the activity and stability of GH7 CBHs is thus of keen interest. Accordingly, we report the comprehensive characterization of the GH7 CBH secreted by G. candidum (GcaCel7A). The bimodular cellulase consists of a family 1 cellulose-binding module (CBM) and linker connected to a GH7 catalytic domain that shares 64% sequence identity with the archetypal industrial GH7 CBH of Hypocrea jecorina (HjeCel7A). GcaCel7A shows activity on Avicel cellulose similar to HjeCel7A, with less product inhibition, but has a lower temperature optimum (50 degrees C versus 60-65 degrees C, respectively). Five crystal structures, with and without bound thio-oligosaccharides, show conformational diversity of tunnel-enclosing loops, including a form with partial tunnel collapse at subsite -4 not reported previously in GH7. Also, the first O-glycosylation site in a GH7 crystal structure is reported - on a loop where the glycan probably influences loop contacts across the active site and interactions with the cellulose surface. The GcaCel7A structures indicate higher loop flexibility than HjeCel7A, in accordance with sequence modifications. However, GcaCel7A retains small fluctuations in molecular simulations, suggesting high processivity and low endo-initiation probability, similar to HjeCel7A.

AB - The ascomycete Geotrichum candidum is a versatile and efficient decay fungus that is involved, for example, in biodeterioration of compact discs; notably, the 3C strain was previously shown to degrade filter paper and cotton more efficiently than several industrial enzyme preparations. Glycoside hydrolase (GH) family 7 cellobiohydrolases (CBHs) are the primary constituents of industrial cellulase cocktails employed in biomass conversion, and feature tunnel-enclosed active sites that enable processive hydrolytic cleavage of cellulose chains. Understanding the structure-function relationships defining the activity and stability of GH7 CBHs is thus of keen interest. Accordingly, we report the comprehensive characterization of the GH7 CBH secreted by G. candidum (GcaCel7A). The bimodular cellulase consists of a family 1 cellulose-binding module (CBM) and linker connected to a GH7 catalytic domain that shares 64% sequence identity with the archetypal industrial GH7 CBH of Hypocrea jecorina (HjeCel7A). GcaCel7A shows activity on Avicel cellulose similar to HjeCel7A, with less product inhibition, but has a lower temperature optimum (50 degrees C versus 60-65 degrees C, respectively). Five crystal structures, with and without bound thio-oligosaccharides, show conformational diversity of tunnel-enclosing loops, including a form with partial tunnel collapse at subsite -4 not reported previously in GH7. Also, the first O-glycosylation site in a GH7 crystal structure is reported - on a loop where the glycan probably influences loop contacts across the active site and interactions with the cellulose surface. The GcaCel7A structures indicate higher loop flexibility than HjeCel7A, in accordance with sequence modifications. However, GcaCel7A retains small fluctuations in molecular simulations, suggesting high processivity and low endo-initiation probability, similar to HjeCel7A.

KW - biomass degradation

KW - cellulase

KW - Geotrichum candidum

KW - molecular dynamics

KW - X-ray structure

KW - TRICHODERMA-REESEI CELLOBIOHYDROLASE

KW - AROMATIC-CARBOHYDRATE INTERACTIONS

KW - PHANEROCHAETE-CHRYSOSPORIUM CEL7D

KW - FAMILY 7 CELLOBIOHYDROLASE

KW - X-RAY

KW - 3-DIMENSIONAL STRUCTURE

KW - TALAROMYCES-EMERSONII

KW - GLYCOSIDE HYDROLASES

KW - ANGSTROM RESOLUTION

KW - BINDING MODULES

U2 - 10.1111/febs.13509

DO - 10.1111/febs.13509

M3 - статья

VL - 282

SP - 4515

EP - 4537

JO - FEBS Journal

JF - FEBS Journal

SN - 1742-464X

IS - 23

ER -