Molecular cloning, overexpression, characterization, and In silico modelling analysis of a novel GDSL autotransporter-dependent outer membrane lipase (OML) of Pseudomonas guariconensis.

The outer membrane lipase (oml) gene, encoding a novel autotransporter-dependent lipase from Pseudomonas guariconensis, was cloned and sequenced. The oml gene has an open reading frame of 1866 bp. It encodes the 621 amino acid autotransporter-dependent GDSL lipase (OML), which has the highest sequence similarity (64.08 %) with the EstA of Pseudomonas aeruginosa (PDB:3kvn.1. A). OML was expressed and purified, which showed a purified band of approximately 70 kDa. The purified enzyme showed maximum activity at pH 9 and 40 °C. Substrate specificity studies and kinetic study by Lineweaver-Burk plot of purified OML showed Km of 1.27 mM and Vmax of 333.33 U/mL with p-nitrophenyl palmitate. The purified enzyme showed good stability in the presence of hexane, methanol, and ethanol, while the presence of the metal ion Mg2+ showed maximum lipase activity. Bioinformatics analysis supported the in vitro findings by predicting enzyme substrate specificity towards long-chain fatty acids and fatty acids with shorter chain lengths. The stability of the interaction of the protein-ligand complex (OML-ricinoleic acid) was confirmed using MDS and castor oil bioconversion using purified OML was confirmed using High-Performance Liquid Chromatography (HPLC).

Analytical and Bioanalytical Techniques for the Quantification of the Calcium Channel Blocker - Amlodipine: A Critical Review.

Hypertension is a condition in which blood pressure is elevated to an extent where benefit is obtained from blood pressure lowering. The risk of complications is proportional to the level that blood pressure raises. Calcium channel blockers are a class of compounds used in the treatment of hypertension. The dihydropyridine (DHP) group, a subclass of the calcium channel blocker works almost exclusively on L-type calcium channels in the peripheral arterioles and reduce blood pressure by reducing total peripheral resistant. Long acting DHP is preferred because they are more convenient for patients and avoid the large fluctuations in plasma drug concentration which are associated with side effects. Amlodipine is the most distinct DHP and the most popular. The drug was patented in the year 1986 and its commercial sale began by 1990. The current article provides a state of art about the analytical and bioanalytical techniques available for the quantification of drug as a single entity and in combined pharmaceutical formulations between 1989 and 2019.

Linezolid-A Review of Analytical Methods in Pharmaceuticals and Biological Matrices.

Bacterial resistance to antibiotics is a growing phenomenon in the world. Considering the relevance of antimicrobials for population and the reduction in the registration of new antimicrobials by regulatory agencies, proper quality control is required to minimize the spread of bacterial resistance and ensure the effectiveness of a treatment, as well as safety for the patient. The recent addition to the antimicrobial world is the oxazolidinone classes of antibiotics, especially useful to treat infections caused by Gram-positive bacteria. Eperezolid and linezolid (LIN) are the two members of the oxazolidinone class of antibiotics. LIN was the first oxazolidinone approved by the Food and Drug Administration. The present review focuses on the analytical methods for the assessment of LIN in pharmaceuticals and biological matrices. The critical validation parameters like the linearity, limit of detection, limit of quantification are discussed for the individual method. Also the critical quality attributes like the sensitivity and the sample preparation techniques for bioanalytical methods are also discussed. Furthermore, some future trends that can be incorporated in the determination of similar drugs are also suggested.

Selective lighting up of segments around Gly, Ala and Ser/Thr in proteins.

Direct detection of (13) C nucleus can be used as a valuable alternative where (1) H detection poses a challenge due to relaxation effects, chemical exchange and poor chemical shift dispersion. In this context, we have designed a suite of 2D (13) C(α) -detected hNCA experiments that provide sequential correlations of (13) C(α) with (15) N on one hand and efficient spectroscopic labeling of certain groups of residues, namely, Gly, Ala, Ser and Thr, on the other. These residues act as checkpoints in the sequential walk, which in turn offer new possibilities of backbone assignment of small proteins from a set of 2D experiments, thereby providing great economy in terms of spectrometer time. The direct identification of peptide segments around Gly, Ala, Ser and Thr residues along a protein chain will be highly valuable for deriving important information on sites of ligand binding, phosphorylation, inhibitor/substrate binding, understanding protein folding pathways, comprehending local conformational dynamics etc. without having to obtain complete sequence-specific assignments, which can be time consuming and at times formidable, especially in large proteins. We have illustratively demonstrated the multifaceted applications of these variants of 2D experiments on ubiquitin and M-crystallin. We foresee that these 2D hNCA experiments will provide economic and efficient strategies for studying the structure and function of proteins.