Penicillin binding proteins (PBPs) have been extensively studied due to their importance to the physiology of bacterial cell wall peptidoglycan and as targets of the most widely used class of antibiotics, the β-lactams. The existing paradigm asserts that PBPs catalyze the final step of peptidoglycan biosynthesis, and β-lactams inhibit their activities. According to this paradigm, a distinct enzyme class, β-lactamases, exists to inactivate β-lactams. This paradigm has been the basis for how bacterial diseases are treated with β-lactams. We tested whether this historical view accurately reflects the relationship between β-lactams and the PBPs and the β-lactamase, BlaC, of Mycobacterium tuberculosis. BlaC was the major inactivator of the cephalosporin subclass of β-lactams. However, the PBPs PonA1 and PonA2 inactivated penicillins and carbapenems more effectively than BlaC. These findings demonstrate that select M. tuberculosis PBPs are effective at inactivating several β-lactams. Lesser-known PBPs, DacB, DacB1, DacB2, and Rv2864c, a putative PBP, were comparably more resistant to inhibition by all β-lactam subclasses. Additionally, Rv1730c exhibited low affinity to most β-lactams. Based on these findings, we conclude that in M. tuberculosis, BlaC is not the only source of inactivation of β-lactams. Therefore, the historical paradigm does not accurately describe the relationship between β-lactams and M. tuberculosis. IMPORTANCE M. tuberculosis, the causative agent of tuberculosis, kills more humans than any other bacterium. β-lactams are the most widely used class of antibiotics to treat bacterial infections. Unlike in the historical model that describes the relationship between β-lactams and M. tuberculosis, we find that M. tuberculosis penicillin binding proteins are able to inactivate select β-lactams with high efficiency.

ABSTRACTThe prevalence of Mycobacteroides abscessus, Mab, (also known as Mycobacterium abscessus) has been increasing steadily globally. Patients with structural lung conditions such as bronchiectasis, cystic fibrosis and chronic obstructive pulmonary disease are at high risk of developing pulmonary Mab disease. The disease is often recurrent as the current treatment regimen is considered sub-efficacious. The cell wall peptidoglycan of bacteria is required for their viability and its biosynthetic pathway is enriched in proteins whose inhibition is the basis for two of the most widely used classes of antibiotics to treat bacterial infections. The peptidoglycan of Mab is distinct from that of most bacteria as its synthesis involves penicillin binding proteins (PBP) and L,D-transpeptidases. Here, we demonstrate that Mab gene locus MAB_3167c encodes a PBP (hereafter referred to as Glby) and is required for normal planktonic growth in liquid broth. Glby exhibits a strong β-lactamase activity and is sensitive to β-lactamase inhibitors. In a mouse model of pulmonary Mab disease, mutant lacking this gene was unable to proliferate, gradually cleared and undetectable after three weeks. In a collection of 1.046 Mab clinical isolates, there is evidence that changes in amino acid sequence that compromise Glby function are not favored. These evidences suggest that an agent that can inhibit Glby in vivo has the potential to be an efficacious treatment against Mab disease.

We present further study of a subset of carbapenems, arising from a previously reported machine learning approach, with regard to their mouse pharmacokinetic profiling and subsequent study in a mouse model of sub-acute Mycobacterium tuberculosis infection. Pharmacokinetic metrics for such small molecules were compared to those for meropenem and biapenem, resulting in the selection of two carbapenems to be assessed for their ability to reduce M . tuberculosis bacterial loads in the lungs of infected mice. The original syntheses of these two carbapenems were optimized to provide multigram quantities of each compound. One of the two experimental carbapenems, JSF-2204, exhibited efficacy equivalent to that of meropenem, while both were inferior to rifampin. The lessons learned in this study point toward the need to further enhance the pharmacokinetic profiles of experimental carbapenems to positively impact in vivo efficacy performance.

Glutathione-S-transferase is one of the important enzyme systems that plays vital role in decomposition of lipid hydro-peroxides formed due to oxidative stress. In the present study GST activity increased in the lymphocytes treated with increasing concentration of H2O2, and decrease in the levels of GSH was observed. For similar treatment conditions LDH activity and MDA levels increased significantly leading to decrease in the cell viability. Treatment of lymphocytes with the root extract of Phlogacanthus tubiflorus (PTE) resulted in dose dependent decline in the GST activity and rise in GSH levels. LDH activity and MDA levels also declined that led to the increase of cell viability. Lymphocytes pre-treated with the PTE followed by H2O2 (0.1 and 1%) treatment, decline in the activity of GST and increase in GSH levels was observed. Also we have observed decline in the activity of LDH and MDA levels in the lymphocytes for both 0.1 and 1% of H2O2 though the magnitude of change was higher in the lymphocytes pre-treated with the PTE followed with 1% of H2O2 treatment. Significant increase in the cell viability for similar conditions was also observed. These findings suggest protective function of the root extracts might be through modulation of GST activity and levels of GSH and might find application in Chemomodulation in future.