Complete genome sequence of carbapenem-resistant pathogenic Klebsiella aerogenes strain CH7 isolated from vermicompost.

We announce the complete genome of Klebsiella aerogenes strain CH7, isolated from a vermicompost sample. A total of 9.14131 million high-quality reads comprised 96 contigs with 5,273 genes and 5,038 protein-coding genes.

Characterization of a Class A ß-Lactamase from Francisella tularensis (Ftu-1) Belonging to a Unique Subclass toward Understanding AMR.

ß-lactamase production with vast catalytic divergence in the pathogenic strain limits the antibiotic spectrum in the clinical environment. Class A carbapenemase shares significant sequence similarities, structural features, and common catalytic mechanisms although their resistance spectrum differs from class A ß-lactamase in carbapenem and monobactam hydrolysis. In other words, it limited the antibiotic treatment option against infection, causing carbapenemase-producing superbugs. Ftu-1 is a class A ß-lactamase expressed by the Francisella tularensis strain, a potent causative organism of tularemia. The chromosomally encoded class A ß-lactamase shares two conserved cysteine residues, a common characteristic of a carbapenemase, and a distinctive class in the phylogenetic tree. Complete biochemical and biophysical characterization of the enzyme was performed to understand the overall stability and environmental requirements to perform optimally. To comprehend the enzyme-drug interaction and its profile toward various chemistries of ß-lactam and ß-lactamase inhibitors, comprehensive kinetic and thermodynamic analyses were conducted using various ß-lactam drugs. The dynamic property of Ftu-1 ß-lactamase was also predicted using molecular dynamics (MD) simulation to compare its loop flexibility and ligand binding with other related class A ß-lactamases. Overall, this study fosters a comprehensive understanding of Ftu-1, proposed to be an intermediate class by characterizing its kinetic profiling, stability by biochemical and biophysical methodologies, and susceptibility profiling. This understanding would be beneficial for the design of new-generation therapeutics.

An insight into omics analysis and metabolic pathway engineering of lignin-degrading enzymes for enhanced lignin valorization.

Lignin, a highly heterogeneous polymer of lignocellulosic biomass, is intricately associated with cellulose and hemicellulose, responsible for its strength and rigidity. Lignin decomposition is carried out through certain enzymes derived from microorganisms to promote the hydrolysis of lignin. Analyzing multi-omics data helps to emphasize the probable value of fungal-produced enzymes to degrade the lignocellulosic material, which provides them an advantage in their ecological niches. This review focuses on lignin biodegrading microorganisms and associated ligninolytic enzymes, including lignin peroxidase, manganese peroxidase, versatile peroxidase, laccase, and dye-decolorizing peroxidase. Further, enzymatic catalysis, lignin biodegradation mechanisms, vital factors responsible for lignin modification and degradation, and the design and selection of practical metabolic pathways are also discussed. Highlights were made on metabolic pathway engineering, different aspects of omics analyses, and its scope and applications to ligninase enzymes. Finally, the advantages and essential steps of successfully applying metabolic engineering and its path forward have been addressed.

Variations in the SDN Loop of Class A Beta-Lactamases: A Study of the Molecular Mechanism of BlaC (Mycobacterium tuberculosis) to Alter the Stability and Catalytic Activity Towards Antibiotic Resistance of MBIs.

The emergence of multidrug-resistant (MDR) and extensively drug-resistant (XDR) tuberculosis calls for an immediate search for novel treatment strategies. Recently, BlaC, the principal beta-lactamase of Mycobacterium tuberculosis, was recognized as a potential therapeutic target. BlaC belongs to Ambler class A, which is generally susceptible to the beta-lactamase inhibitors currently used in clinics: tazobactam, sulbactam, and clavulanate. Alterations at Ser130 in conserved SDN loop confer resistance to mechanism-based inhibitors (MBIs) commonly observed in various clinical isolates. The absence of clinical evidence of S130G conversion in M. tuberculosis draws our attention to build laboratory mutants of S130G and S130A of BlaC. The study involving steady state, inhibition kinetics, and fluorescence microscopy shows the emergence of resistance against MBIs to the mutants expressing S130G and S130A. To understand the molecular reasoning behind the unavailability of such mutation in real life, we have used circular dichroism (CD) spectroscopy, differential scanning calorimetry (DSC), molecular dynamics (MD) simulation, and stability-based enzyme activity to compare the stability and dynamic behaviors of native and S130G/A mutant form of BlaC. A significant decrease in melting temperature (BlaC T M 60°C, S130A T M 50°C, and S130G T M 45°C), kinetic instability at higher temperature, and comparative dynamic instability correlate the fact that resistance to beta-lactam/beta-lactamase inhibitor combinations will likely not arise from the structural alteration of BlaC, therefore establishing confidence that this therapeutic modality can be potentially applied as a part of a successful treatment regimen against M. tuberculosis.

An insight into the complete biophysical and biochemical characterization of novel class A beta-lactamase (Bla1) from Bacillus anthracis.

Bacillus anthracis, a potent pathogen of anthrax is becoming resistant to many beta-lactam antibiotics because of the expression of two chromosomally encoded beta-lactamases Bla1 and Bla2. Bla1 is a class A beta-lactamase whereas Bla2 is a Metallo beta-lactamase. In the current study, we have attempted in-detailed characterization of Bla1 beta-lactamase by taking interdisciplinary approaches. Our study includes structure and sequence comparison of this enzyme with other members of the class, to know the conservation pattern that includes active site residues, secondary structure, conserved fold, evolutionary relationships, etc. Dynamic characterizations of the enzyme, unfolding kinetics were determined with the help of Molecular dynamics simulation. Detailed enzyme stability and catalytic activity towards various physical (Temperature and pH), and chemical parameters (Urea, GnHCl) were performed. Together, our study helps to develop a proper understanding of this beta-lactamase by characterizing its biochemical, biophysical, dynamic, kinetic and thermodynamic properties. This would help contribute towards a better understanding of beta-lactamase based AMR emergence.