Production of mycobacterial cell wall glycopeptidolipids requires a member of the MbtH-like protein family

ABSTRACT: BACKGROUND: Glycopeptidolipids (GPLs) are among the major free glycolipid components of the outer membrane of several saprophytic and clinically-relevant Mycobacterium species. The architecture of GPLs is based on a constant tripeptide-amino alcohol core of nonribosomal peptide synthetase origin that is N-acylated with a 3-hydroxy/methoxy acyl chain synthesized by a polyketide synthase and further decorated with variable glycosylation patterns built from methylated and acetylated sugars. GPLs have been implicated in many aspects of mycobacterial biology, thus highlighting the significance of gaining an understanding of their biosynthesis. Our bioinformatics analysis revealed that every GPL biosynthetic gene cluster known to date contains a gene (referred herein to as gplH) encoding a member of the MbtH-like protein family. Herein, we sought to conclusively establish whether gplH was required for GPL production. RESULTS: Deletion of gplH, a gene clustered with nonribosomal peptide synthetase-encoding genes in the GPL biosynthetic gene cluster of Mycobacterium smegmatis, produced a GPL deficient mutant. Transformation of this mutant with a plasmid expressing gplH restored GPL production. Complementation was also achieved by plasmid-based constitutive expression of mbtH, a paralog of gplH found in the biosynthetic gene cluster for production of the siderophore mycobactin of M. smegmatis. Further characterization of the gplH mutant indicated that it also displayed atypical colony morphology, lack of sliding motility, altered capacity for biofilm formation, and increased drug susceptibility. CONCLUSIONS: Herein, we provide evidence formally establishing that gplH is essential for GPL production in M. smegmatis. Inactivation of gplH also leads to a pleiotropic phenotype likely to arise from alterations in the cell envelope due to the lack of GPLs. While genes encoding MbtH-like proteins have been shown to be needed for production of siderophores and antibiotics, our study presents the first case of one such a gene proven to be required for production of a cell wall component. Furthermore, our results provide the first example of a mbtH-like gene with confirmed functional role in a member of the Mycobacterium genus. Altogether, our findings demonstrate a critical role of gplH in mycobacterial biology and advance our understanding of the genetic requirements for the biosynthesis of an important group of constituents of the mycobacterial outer membrane.     

Occult hepatitis C virus infection in patients with beta-thalassemia major: Is it a neglected and unexplained phenomenon?

Occult hepatitis C virus (HCV) infection (OCI) is described as the presence of viral genome in both hepatocytes and peripheral blood mononuclear cells (PBMCs) despite constant negative results on serum HCV RNA tests. Beta-thalassemia major (BTM) describes a group of inherited blood diseases. Patients with BTM require repeated blood transfusions, increasing the risk of exposure to infectious agents. We aimed to assess the prevalence of OCI in Iranian BTM patients and to identify the role of host factors in OCI positivity. A total of 181 BTM patients with HCV negative markers were selected. HCV RNA was tested in PBMCs using nested polymerase chain reaction assay. The positive samples were then genotyped via restriction fragment-length polymorphism (RFLP) and 5′-untranslated region sequencing. Six (3.3%) out of 181 BTM patients had viral HCV genomes in PBMC samples. Three (50.0%), two (33.3%), and one (16.7%) out of these six patients were infected with HCV-1b, HCV-1a, and HCV-3a, respectively. OCI positivity was significantly associated with the serum level of uric acid (P = 0.045) and ABO blood group (P = 0.032). Also, OCI patients had unfavorable IFNL3 rs12979860 TT, IFNL3 rs8099917 GG, IFNL3 rs12980275 GG, and IFNL4 ss469415590 ∆G/∆G genotypes. In conclusion, we indicated the low frequency of OCI in BTM patients. Nevertheless, more attention is warranted considering the importance of this infection. Also, further studies are necessary to determine the actual prevalence of OCI among BTM patients in Iran.     

Rich can get poor: conversion of hub to non-hub proteins

Hubs are ubiquitous network elements with high connectivity. One of the common observations about hub proteins is their preferential attachment leading to scale-free network topology. Here we examine the question: does rich protein always get richer, or can it get poor too? To answer this question, we compared similar and well-annotated hub proteins in six organisms, from prokaryotes to eukaryotes. Our findings indicate that hub proteins retain, gain or lose connectivity based on the context. Furthermore, the loss or gain of connectivity appears to correlate with the functional role of the protein in a given system.     

Index to volume 8 subject index

The purpose of this study was to determine if changes in peripheral levels of E or F series prostaglandin (PGE or PGF) during pseudopregnancy (PSP) in the rat can be correlated with the changes in peripheral levels of total progestin (Total P), and if estrogen surge on day 4 is associated with increased levels of PGE or PGF. The results indicate that an increase in the concentration of PGF on day 7 may have precipitated a gradual decline in peripheral P. However, no correlation was detected between PGE and peripheral total P. Furthermore, no preimplantation increase in PGE or PGF levels was detected, even though the concentrations of these PG's were relatively high during the first 4 days of PSP.     

Design,Synthesis, Molecular Docking,ADMET Studies,and Biological Evaluation of Isoxazoline and Pyrazoline Incorporating 1,2,3-Triazole Benzene Sulfonamides

Russian Journal of Bioorganic Chemistry - In targeted therapy of cancer, PI3Kα targeting is being considered as a promising approach to design novel anticancer drugs. In the present work we...     

Urinary Prognostic Biomarkers and Classification of IgA Nephropathy by High Resolution Mass Spectrometry Coupled with Liquid Chromatography

IgA nephropathy is the most common cause of primary glomerulonephritis. There are different pathologic biopsy-based scoring systems in use, but there is no consensus among nephrologists yet regarding the best classification method. Our aim was to test urine proteomics as a non-invasive method for classification of IgA nephropathy. This aim was pursued by discovering novel prognostic protein biomarkers in urine, and linking them to pathogenesis of the disease through known signaling and metabolic pathways. 13 urine samples of the patients with biopsy-proven IgA nephropathy were analyzed via two proteomics approaches: nanoflow LC-MS/MS and GeLC-MS/MS. The results of label-free quantification were subjected to multivariate statistical analysis, which could classify patients into two groups, broadly corresponding to the primary and advance stages. The proteome classification correlated well with biopsy-based scoring systems, especially endocapillary hypercellularity score of the Oxford’s classification. Differentially excreted candidate proteins were found as potential prognostic biomarkers: afamin, leucine-rich alpha-2-glycoprotein, ceruloplasmin, alpha-1-microgolbulin, hemopexin, apolipoprotein A-I, complement C3, vitamin D-binding protein, beta-2-microglobulin, and retinol-binding protein 4. Pathway analysis suggested impairment of Extra Cellular Matrix (ECM)-Receptor Interaction pathways as well as activation of complement and coagulation pathway in progression of IgA nephropathy.     

Study of the mechanism of interaction of antibody (IgG) on two mixed mode sorbents

Purification of target proteins from a crude biological mixture containing proteins, peptides and other biomolecules is the chromatographic challenge. Mixed mode chromatography offers additional selectivities to improve the overall productivity of commercial bioprocesses with novel chromatographic sorbents being introduced to overcome the problem. HEA HyperCel? (n-hexyl amine) and PPA HyperCel? (phenyl propyl amine) are industry scalable mixed mode chromatography sorbents where both hydrophobic and electrostatic interactions are predominant. Our study focuses on understanding the underlying mechanism of interaction of protein with the sorbent. Parameters like buffer conditions, pH and temperature were tuned to study the adsorption and desorption conditions of the protein. Dynamic binding capacity of HEA HyperCel? and PPA HyperCel? sorbents was studied with human IgG as a model protein. Our study shows that, in HEA the interaction of IgG to the sorbent is predominantly hydrophobic as the binding is enhanced (50–60 mg/ml of sorbent) by presence of salt in buffer and increase in temperature. Binding capacity of PPA is 50–60 mg/ml of sorbent irrespective of temperature effect and/or the presence of salt. The chromatographic experiments show that the interaction could be hydrophobic or ionic or some charge transfer mechanism depending upon the buffer conditions.     

Caste and sex specific olfactory glomerular organization and brain architecture in two sympatric ant species Camponotus sericeus and Camponotus compressus (Fabricius, 1798)

We use monoclonal antibodies against synaptic proteins and anterograde tracing with neurobiotin to describe the architecture of the antennal lobes in different castes of two ant species – Camponotus sericeus and Camponotus compressus. The reproductives and worker classes are readily categorized based on size and external morphology. The overall organization of brain neuropile is comparable between castes with differences only in the visual ganglia. Males have a larger fraction of neuropile occupied by the medulla and lobula than females. In the diurnal species, C. sericeus these regions are more highly represented, than in the nocturnal species C. compressus. The most striking differences are in the antennal lobe where males possess a macroglomerulus, which is about ten times larger in volume than the other glomeruli; such a specialization is absent in females. Minor workers possess a significantly larger number of glomeruli than the majors despite the smaller overall volume of the lobe. These caste-specific differences occur mainly within glomerular clusters that receive input from sensory neurons that project in tracts – T4 and T5 – within the antennal nerve. The comparative anatomy of different castes of ants provides an entry point into a future systematic analysis of how divergent brain architectures can arise within a single species.     

Design and Synthesis of Heterocyclic Conjugated Peptides as Novel Antimicrobial Agents

Antimicrobial peptides have been recognized as a novel class of antibiotics and several candidates are currently in clinical trials. In the present study, new antimicrobial compounds were synthesized by coupling quinazolinone moiety with the fragments of elastin sequences VP, GVP, VGVP and GVGVP. They were evaluated for their antibacterial activity against both gram positive and gram negative bacterial strains. We are here reporting that heterocyclic conjugated tetra peptide and penta peptide showed enhanced antibacterial activity compare to the conventional antimicrobial drugs.     

Proline accumulation and the expression of Δ1-pyrroline-5-carboxylate synthetase in two safflower cultivars

Proline (Pro) accumulation under water stress was measured in safflower (Carthamus tinctorius L.) drought tolerant cv. A1 and sensitive cv. Nira. Activities of pyrroline-5-carboxylate reductase (P5C reductase) and pyrroline-5-carboxylate synthetase (P5C synthetase), two enzymes involved in the Pro biosynthetic pathway were also estimated. Water stress resulted in a reduction in the leaf dry mass and chlorophyll content along with a gradual accumulation of Pro. RT-PCR results show higher expression of Δ¹-pyrroline-5-carboxylate synthetase (p5cs) gene in correlation with up-regulated Pro accumulation in cv. A1. P5C reductase was found to be the Pro synthesis rate limiting whereas P5C synthetase did not show any specific response to the drought stress in both cultivars.