Biology subjects

Panda, A. P.

Publications and source records attributed to Panda, A. P..

4 recordsLinked to original sources

bioRxiv2026

Rv0783c of Mycobacterium tuberculosis acts as a proton-motive force dependent multidrug efflux transporter involved in the efflux of structurally unrelated antibiotics and enhancing biofilm formation

Despite multiple treatment strategies and extensive research on resistance mechanisms, tuberculosis (TB) remains a major global health threat, largely because of the rise of multidrug-resistant (MDR) and extensively drug-resistant (XDR) TB. Among various mechanisms complicating the situation, active antibiotic export via efflux pumps is particularly significant, yet largely unexplored. Mycobacterium sp. encodes numerous transporters, many of which are overexpressed in clinical isolates or under drug stress. Here, we examined the possible role of Rv0783c, a putative transporter that is reportedly overexpressed in drug-stressed conditions. Rv0783c conferred resistance to multiple structurally diverse antibiotics, fluoroquinolones and anti-TB drugs in the heterologous hosts, namely, Escherichia coli and Mycobacterium smegmatis. Reduced drug accumulation and active efflux of ethidium bromide (EtBr) confirmed its transport activity, which in turn gets nullified upon using the proton-motive force blocker, CCCP. On the other hand, its expression enhanced biofilm formation, linking antibiotic resistance to persistence-associated phenotype. Furthermore, site-directed mutagenesis confirmed the presence of crucial interacting residues with antibiotics that were identified by in silico analysis. Overall, we demonstrate the role of Rv0783c in the extrusion of first and second-line anti-TB drugs and enhancing biofilm formation.

microbiology↗

bioRxiv2025

Substitution of a valine to glutamic acid in the omega-like loop of MSMEG_6194 of Mycobacterium smegmatis interchanges its activity from DD-carboxypeptidase to beta-lactamase

The genome of Mycobacterium smegmatis encodes numerous penicillin-interacting enzymes, and analysing their functions provides insights into the evolutionary mechanisms behind beta-lactam resistance in mycobacteria. In this study, we characterised one such enzyme, MSMEG_6194, annotated as a putative beta-lactamase. Although MSMEG_6194 shares structural similarity with class A beta-lactamases, it showed no detectable beta-lactamase activity under the tested conditions. Heterologous expression of MSMEG_6194 in Escherichia coli and{Delta} msmeg_6194 deleted strains of M. smegmatis did not confer significant resistance to beta-lactams, and the purified protein failed to hydrolyse nitrocefin either. However, ectopic expression of MSMEG_6194 partly restores the morphological defects in seven PBP-deleted E. coli strains, and the purified enzyme successfully cleaves the terminal D-alanine from a pentapeptide substrate, confirming its DD-carboxypeptidase activity. Structural analysis revealed the absence of a conserved glutamic acid residue in the omega-loop, which is critical for beta-lactamase catalysis in class A beta-lactamase. Substituting this residue (V139E mutant) imparts beta-lactamase activity though significantly reduces DD-carboxypeptidase function. Overall, these findings establish MSMEG_6194 as a DD-carboxypeptidase and demonstrate how a single amino acid change can alter catalytic preference, shedding light on the evolutionary transition from DD-Carboxypeptidases to beta-lactamases in mycobacteria.

molecular biology↗

bioRxiv2025

Putative MFS transporter Rv1250 of Mycobacterium tuberculosis is involved in multidrug efflux activity

Drug-resistant Mycobacterium tuberculosis is one of the leading causes of global mortality. Mechanisms, such as slow uptake of drugs along with cell wall impermeability and active efflux, are some of the concerning reasons leading to drug resistance. Efflux pumps actively transport a wide variety of drugs and toxins away from the target site, which is considered an emerging cause for the failure of anti-tubercular medications and treatment. In this study, we report that the ability of Rv1250, a probable MFS-type transporter, influences the extrusion of multiple structurally unrelated classes of drugs, enhances the biofilm formation in E. coli and Mycobacterium smegmatis, and facilitates the survival of M. smegmatis cells inside the macrophage during antibiotic stress. Interestingly, in trans, the expression of rv1250 decreased the susceptibility of host cells to several structurally unrelated antibiotics, ranging from fluoroquinolones to aminoglycosides, beta-lactams, and anti-tubercular drugs, thus indicating its involvement in imparting intrinsic drug tolerance. In addition, the increased efflux of EtBr, norfloxacin, and Bocillin FL from host cells expressing rv1250 was revealed by the hosts ability to confer a lower level of antibiotic accumulation. Moreover, the expression of rv1250 resulted in the enhancement of biofilm formation. Overall, we conclude that Rv1250 of Mycobacterium tuberculosis might facilitate the survival of host cells under antimicrobial stress. One sentence summaryRole of Rv1250 as an efflux pump

microbiology↗

bioRxiv2024

A magnesium transporter CorA of Mycobacterium smegmatis enhances the tolerance of structurally unrelated antibiotics in the host cells

Ion transporters or channels are involved in maintaining metal homeostasis in bacterial cells by aiding the movement of metal ions across the cell, which might also facilitate the export of antimicrobials. Ubiquitous magnesium transporter, CorA of Mycobacterium smegmatis is well known for its role in maintaining magnesium homeostasis. However, little is known about its involvement in exerting antimicrobial resistance. Here, with the help of molecular genetics, in vivo and in silico studies we tried to envisage the role of CorA of M. smegmatis in antimicrobial resistance of M. smegmatis and E. coli. Expression of corA in M. smegmatis and E. coli increased the tolerance of the host cells towards various structurally unrelated antibiotics and anti-tubercular drugs. In addition, a significantly lower accumulation of norfloxacin and ofloxacin by the host cells expressing corA further indicated its role in enhancing the efflux pump activity. Moreover, the presence of a sub-inhibitory concentration of Mg2+ resulted in increased low-level tolerance towards the tested drugs. Furthermore, CorA enhanced the biofilm-forming ability of cells expressing it. Overall, we speculate that magnesium transporter CorA facilitates multi-drug efflux activity of the host cells where Mg2+ might act as a facilitator in the process. IMPORTANCEMagnesium acts as a co-factor for various biochemical and physiological reactions, such as protein synthesis, cell membrane integrity, nucleic acid synthesis, etc. Metal transporters maintain metal homeostasis by regulating the uptake, efflux, or transportation of metals in certain necessary cellular compartments. In bacteria, magnesium ion (Mg2+) is mainly supplied by the CorA protein which is a ubiquitous family of transport proteins and extensively studied in E. coli and Salmonella sp. However, little is known about the functional relationship of metal transporters of Mycobacterium sp with extrusion of antibiotics, and their involvement in stress tolerance. Here, we report CorA (MSMEG_5056), a magnesium transporter of Mycobacterium smegmatis in influencing the extrusion of multiple structurally unrelated classes of drugs and enhancing the biofilm formation of E. coli and Mycobacterium smegmatis. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=74 SRC="FIGDIR/small/602764v1_ufig1.gif" ALT="Figure 1"> View larger version (26K): org.highwire.dtl.DTLVardef@11a5f5corg.highwire.dtl.DTLVardef@6ac8f9org.highwire.dtl.DTLVardef@b083dcorg.highwire.dtl.DTLVardef@a0fb8a_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOGraphical Abstract:C_FLOATNO Hypothetical model of Antibiotic export by CorA. Antibiotics bind to the closed state of the protein (left). During the transition to the open state (right), the to-and-fro motion between multiple open states drives the efflux of the antibiotic while facilitating the import of Mg2+. The colors of the models correspond to the chain ID. The bottom views of both the closed and open states are shown in the rectangular box, with the color indicated by their respective chain IDs. C_FIG

microbiology↗