The Carbohydrate Active Enzyme(CaZY) database indicates that glycoside hydrolase family 55 (GH55) contains both endo- and exo-β -1,3-glucanases.We present high resolution crystal structures of bacterial SacteLam55A from the highly cellulolytic Streptomyces sp. SirexAA-E with bound substrates and product. These structures, along with mutagenesis and kinetic studies implicate Glu502 as the catalytic acid and a proton relay network of four residues in activating water as the nucleophile. Further, a set of conserved aromatic residues that define the active site apparently enforce an exo-glucanase reactivity as demonstrated by exhaustive hydrolysis reactions with purified laminarioligosaccharides. Two additional aromatic residues that line the substratebinding channel show substrate-dependent conformational flexibility that may promote processive reactivity of the bound oligosaccharide in the bacterial enzymes.
Function
SacteLam55A is a GH55 enzyme from highly cellulolytic Streptomyces sp. SirexAA-E. Substrate bound structures identify residues involved in binding, catalysis, enforcement of reaction specificity and possibly processivity. Natural GH55 are exo-β-1,3- glucanases with a broad range of temperature and
pH optima.
Relevance
Experimental annotation of GH phylogenetic space by use of bioinformatics, highthroughput cell-free translation, biochemical assay, and structure determination is feasible.
Structural highlights
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References
Christopher M. Bianchetti, Taichi E. Takasuka, Sam Deutsch, Hannah S. Udell, Eric J. Yik, Lai F. Bergeman, and Brian G. Fox.
