Mycobacterium tuberculosis H37Rv enolase (Rv1023)- expression, characterization and effect of host dependent modifications on protein functionality

Mycobacterium tuberculosis enolase is an essential glycolytic enzyme that catalyzes the conversion of 2, phosphoglycerate (PGA) to phosphoenol pyruvate (PEP). It is also a crucial link between glycolysis and the tricarboxylic acid (TCA) pathway. The depletion of PEP has recently been associated with the emergence of non-replicating drug resistant bacteria. Enolase is also known to exhibit multiple alternate functions, such as promoting tissue invasion via its role as a plasminogen (Plg) receptor. In addition, proteomic studies have identified the presence of enolase in the Mtb degradosome and in biofilms. However, the precise role in these processes has not been elaborated. The enzyme was recently identified as a target for 2-amino thiazoles - a novel class of anti-mycobacterials. In vitro assays and characterization of this enzyme were unsuccessful due to the inability to obtain functional recombinant protein. In the present study, we report the expression and characterization of enolase using Mtb H37Ra as a host strain. Our study demonstrates that the enzyme activity and alternate functions of this protein are significantly impacted by the choice of expression host (Mtb H37Ra or E. coli). Detailed analysis of the protein from each source revealed subtle differences in the post-translational modifications. Lastly, our study confirms the role of enolase in Mtb biofilm formation and describes the potential for inhibiting this process. •Mycobacterium tuberculosis Enolase is a key enzyme linking the glycolytic and TCA pathways.•Here we demonstrate the successful expression of functional Mtb Enolase using Mtb H37Ra as a surrogate.•Recombinant protein obtained from Mtb H37Ra demonstrated increased yield and functionality as compared to the E.coli host.•The enzymes differed in their activity, alternate functions and post-translational modifications.•Lastly, we confirmed the role of Mtb Enolase in biofilm formation.