Distinct functional properties of secretory l-asparaginase Rv1538c involved in phagosomal survival of Mycobacterium tuberculosis.

Abstract

The emergence of drug-resistant Mycobacterium tuberculosis (Mtb) stains has escalated the need for developing more efficient drugs and therapeutic strategies against tuberculosis. Here we functionally annotate a secretory mycobacterial asparaginase Rv1538c (MtA) and describe its biochemical properties. MtA primarily existed as dimer along with a minor population of multimers. Circular dichroism and fluorescence spectroscopy demonstrated a compact structure in Tris HCl buffer at pH 8.0. Under these conditions it also displayed optimum activity. It retained ∼40% activity at pH 5.5, supporting its physiological relevance in acidic phagosomal environment. MtA contravened classical Michaelis-Menten kinetics and exhibited product inhibition profile, yielding a Kcat of 869.4 s-1 and an apparent Km of 8.36\u202fmM. We report the presence of several antigenic epitopes and a C-terminal YXXXD/E motif in MtA, hinting towards its potential to interact or influence host immune system. This was supported by our observation of morphological changes in MtA-treated human B lymphoblasts. We propose that MtA is a dual purpose enzyme used by Mtb to survive inside its host by; 1) ammonia-mediated neutralization of the phagosomal acidic pH and 2) inducing stress to primary immune cells and compromising the host immune response. Overall, this study contributes to our understanding of the biological role of mycobacterial asparaginase opening avenues for developing effective TB therapeutics.