Structure, computational and biochemical analysis of PcCel45A endoglucanase from Phanerochaete chrysosporium and catalytic mechanisms of GH45 subfamily C members

Scientific Reports
Andre S GodoyIgor Polikarpov

Abstract

The glycoside hydrolase family 45 (GH45) of carbohydrate modifying enzymes is mostly comprised of β-1,4-endoglucanases. Significant diversity between the GH45 members has prompted the division of this family into three subfamilies: A, B and C, which may differ in terms of the mechanism, general architecture, substrate binding and cleavage. Here, we use a combination of X-ray crystallography, bioinformatics, enzymatic assays, molecular dynamics simulations and site-directed mutagenesis experiments to characterize the structure, substrate binding and enzymatic specificity of the GH45 subfamily C endoglucanase from Phanerochaete chrysosporium (PcCel45A). We investigated the role played by different residues in the binding of the enzyme to cellulose oligomers of different lengths and examined the structural characteristics and dynamics of PcCel45A that make subfamily C so dissimilar to other members of the GH45 family. Due to the structural similarity shared between PcCel45A and domain I of expansins, comparative analysis of their substrate binding was also carried out. Our bioinformatics sequence analyses revealed that the hydrolysis mechanisms in GH45 subfamily C is not restricted to use of the imidic asparagine as a general base...Continue Reading

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Citations

Feb 15, 2020·The Journal of Biological Chemistry·Vivek S BharadwajMichael F Crowley
Mar 25, 2021·Journal of Chemical Information and Modeling·Caroline S PereiraMunir S Skaf
Apr 24, 2021·International Journal of Biological Macromolecules·Amisha Patel, Amita Shah
Dec 11, 2021·Applied Microbiology and Biotechnology·Ruiqin ZhangYanjun Li

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Datasets Mentioned

BETA
BAG68300

Methods Mentioned

BETA
X-ray
flow filtration

Software Mentioned

HHPred
OligoDesigner
MEGA7
Clustal Omega
NAMD
BLAST
HTP
PyMOL
DALI
Dali server

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