Comparative analysis of homologous aminopeptidase PepN from Mycobacterium tuberculosis and Mycobacterium smegmatis reveals dichotomy of traits

ABSTRACT: Bacterial pathogens exploit secreted aminopeptidases to modulate host cellular functions. Mycobacterium tuberculosis (Mtb) secretes PepN, a M1 aminopeptidase with N-terminal peptidase and C-terminal ERAP1_C-like domains. In addition to being conserved across Mtb complex, PepNMtb is also homologous (78% amino acids identity) to PepN of M. smegmatis (Msmeg), a non-pathogenic mycobacterium. Despite high homogeneity, PepNMtb and PepNMsmeg exhibit opposing traits thus providing insights into their plausible bacterial/host-specific functions. Our biochemical studies show that, while both PepNs uniformly accumulate across all stages of in vitro growth, unlike Mtb, Msmeg is intolerant to over accumulation of its PepN and hence robustly proteolyzes it. Our proteomic analyses indicate Msmeg also secreting some of its excess PepN into its culture supernatant. In contrast, Mtb does not excessively secrete or proteolyse its over accumulating PepNMtb. Our fractionation studies show Mtb translocating its full length PepNMtb into its membrane and cell wall fractions. Instead, Msmeg accumulates its full length PepNMsmeg in its cytosol, and promotes its cleaved (~40 kDa) form to locate into subcellular fractions. PepNMtb is redundant for Mtb’s in vitro growth and thus Mtb is readily amenable to ΔpepN generation. Opposingly, PepNMsmeg is necessary but not sufficient for Msmeg growth in vitro. Given such disparity, co-immunoprecipitation studies on both PepNs predictably showed divergent interactomes with minimal common hits. In our in vitro infection studies, PepNMtb localizes host endoplasmic reticulum directing us to its potential host-specific role. In summary, our study provides insights into PepNs dichotomy despite them being conserved across pathogenic and non-pathogenic mycobacterial species.

INSTRUMENT(S): Q Exactive Plus

ORGANISM(S): Mycobacterium Tuberculosis Mycobacterium Smegmatis

DISEASE(S): Tuberculosis

SUBMITTER: Saravanan Kumar  

LAB HEAD: Krishnamohan Atmakuri

PROVIDER: PXD009164 | Pride | 2019-07-29

REPOSITORIES: Pride