PDBsum entry 5go5

Hydrolase

PDB id: 5go5

Contents

Protein chain

152 a.a.

Ligands

GLY

SO4 ×2

Waters ×154

References listed in PDB file

Key reference

• Title: Structure and specificity of a new class of ca²⁺-Independent housekeeping sortase fromstreptomyces avermitilisprovide insights into its non-Canonical substrate preference.
• Authors: S.Das, V.S.Pawale, V.Dadireddy, A.K.Singh, S.Ramakumar, R.P.Roy.
• Ref.: J Biol Chem, 2017, 292, 7244-7257.
• PubMed id: 28270507

Abstract

Surface proteins in Gram-positive bacteria are incorporated into the cell wall through a peptide ligation reaction catalyzed by transpeptidase sortase. Six main classes (A-F) of sortase have been identified of which class A sortase is meant for housekeeping functions. The prototypic housekeeping sortase A (SaSrtA) fromStaphylococcus aureuscleaves LPXTG-containing proteins at the scissile T-G peptide bond and ligates protein-LPXT to the terminal Gly residue of the nascent cross-bridge of peptidoglycan lipid II precursor. Sortase-mediated ligation ("sortagging") of LPXTG-containing substrates and Gly-terminated nucleophiles occursin vitroas well asin celluloin the presence of Ca²⁺and has been applied extensively for protein conjugations. Although the majority of applications emanate from SaSrtA, low catalytic efficiency, LPXTG specificity restriction, and Ca²⁺requirement (particularly forin celluloapplications) remain a drawback. Given that Gram-positive bacteria genomes encode a variety of sortases, natural sortase mining can be a viable complementary approach akin to engineering of wild-type SaSrtA. Here, we describe the structure and specificity of a new class E sortase (SavSrtE) annotated to perform housekeeping roles inStreptomyces avermitilisBiochemical experiments define the attributes of an optimum peptide substrate, demonstrate Ca²⁺-independent activity, and provide insights about contrasting functional characteristics of SavSrtE and SaSrtA. Crystal structure, substrate docking, and mutagenesis experiments have identified a critical residue that dictates the preference for a non-canonical LAXTG recognition motif over LPXTG. These results have implications for rational tailoring of substrate tolerance in sortases. Besides, Ca²⁺-independent orthogonal specificity of SavSrtE is likely to expand the sortagging toolkit.