The structure of Rv3717 reveals a novel amidase from Mycobacterium tuberculosis.

Atul Kumar,Sanjiv Kumar,Rikeshwer P Dewangan,Srinivasan Ramachandran,Arpit Mishra,Priyanka Shrivastava,Dilip Kumar,Bhupesh Taneja

doi:10.1107/s0907444913026371

Abstract

Bacterial N-acetylmuramoyl-L-alanine amidases are cell-wall hydrolases that hydrolyze the bond between N-acetylmuramic acid and L-alanine in cell-wall glycopeptides. Rv3717 of Mycobacterium tuberculosis has been identified as a unique autolysin that lacks a cell-wall-binding domain (CBD) and its structure has been determined to 1.7 Å resolution by the Pt-SAD phasing method. Rv3717 possesses an α/β-fold and is a zinc-dependent hydrolase. The structure reveals a short flexible hairpin turn that partially occludes the active site and may be involved in autoregulation. This type of autoregulation of activity of PG hydrolases has been observed in Bartonella henselae amidase (AmiB) and may be a general mechanism used by some of the redundant amidases to regulate cell-wall hydrolase activity in bacteria. Rv3717 utilizes its net positive charge for substrate binding and exhibits activity towards a broad spectrum of substrate cell walls. The enzymatic activity of Rv3717 was confirmed by isolation and identification of its enzymatic products by LC/MS. These studies indicate that Rv3717, an N-acetylmuramoyl-L-alanine amidase from M. tuberculosis, represents a new family of lytic amidases that do not have a separate CBD and are regulated conformationally.

Highlights

Mycobacterial diseases such as tuberculosis and leprosy remain a great threat to human life
Each of these searches identified two N-acetylresearch papers muramoyl-l-alanine amidases in the M. tuberculosis genome: Rv3915, previously characterized as an N-acetylmuramoyl-lalanine amidase (Deng et al, 2005), and Rv3717, previously annotated as a conserved hypothetical protein
Conformational changes in Rv3717 of PlyL to be catalytically active has been postulated to be a upon substrate binding may lead to changes in the surface to consequence of the net positive charge of Z = +6 on the enable more residues to interact with the substrate cell wall protein

Summary

Introduction

Mycobacterial diseases such as tuberculosis and leprosy remain a great threat to human life. An increase in multidrug-resistant cases in both developed and developing countries in recent years, including associated mortality in HIV co-infected patients (World Health Organization, 2011), has resulted in renewed concerted efforts to search for new drugs and drug targets to combat this pathogen (Raviglione et al, 2012; Bhardwaj et al, 2011). The outer lipid layer is esterified with mycolic acids that are covalently linked to the arabinogalactan layer (Hoffmann et al, 2008; Zuber et al, 2008). This arabinogalactan layer is further enclosed by a capsular layer of glycans, lipids and Received 17 July 2013 Accepted 24 September 2013

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