Metabolite Damage and Damage-Control in a Minimal Genome

Analysis of the genes retained in the minimized Mycoplasma JCVI-Syn3A genome established that systems that repair or preempt metabolite damage are essential to life. Several genes with known metabolite damage repair or preemption functions were identified and experimentally validated, including 5-formyltetrahydrofolate cyclo-ligase, CoA disulfide reductase, and certain hydrolases. Furthermore, we discovered that an enigmatic YqeK hydrolase domain fused to NadD has a novel proofreading function in NAD synthesis and could double as a MutT-like sanitizing enzyme for the nucleotide pool. Finally, we combined metabolomics and cheminformatics approaches to extend the core metabolic map of JCVI-Syn3A to include promiscuous enzymatic reactions and spontaneous side reactions. This extension revealed that several key metabolite damage-control systems remain to be identified in JCVI-Syn3A, such as that for methylglyoxal.

Ca2+ imbalance caused by ERdj5 deletion affects mitochondrial fragmentation

The endoplasmic reticulum (ER) is the organelle responsible for the folding of secretory/membrane proteins and acts as a dynamic calcium ion (Ca2+) store involved in various cellular signalling pathways. Previously, we reported that the ER-resident disulfide reductase ERdj5 is involved in the ER-associated degradation (ERAD) of misfolded proteins in the ER and the activation of SERCA2b, a Ca2+ pump on the ER membrane. These results highlighted the importance of the regulation of redox activity in both Ca2+ and protein homeostasis in the ER. Here, we show that the deletion of ERdj5 causes an imbalance in intracellular Ca2+ homeostasis, the activation of Drp1, a cytosolic GTPase involved in mitochondrial fission, and finally the aberrant fragmentation of mitochondria, which affects cell viability as well as phenotype with features of cellular senescence. Thus, ERdj5-mediated regulation of intracellular Ca2+ is essential for the maintenance of mitochondrial homeostasis involved in cellular senescence.

Small Molecule–Peptide Conjugates as Dimerization Inhibitors of Leishmania infantum Trypanothione Disulfide Reductase

Trypanothione disulfide reductase (TryR) is an essential homodimeric enzyme of trypanosomatid parasites that has been validated as a drug target to fight human infections. Using peptides and peptidomimetics, we previously obtained proof of concept that disrupting protein–protein interactions at the dimer interface of Leishmania infantum TryR (LiTryR) offered an innovative and so far unexploited opportunity for the development of novel antileishmanial agents. Now, we show that linking our previous peptide prototype TRL38 to selected hydrophobic moieties provides a novel series of small-molecule–peptide conjugates that behave as good inhibitors of both LiTryR activity and dimerization.

A novel mycothiol-dependent thiol–disulfide reductase in Corynebacterium glutamicum involving oxidative stress resistance

Abstractncgl2478 gene from Corynebacterium glutamicum encodes a thiol–disulfide oxidoreductase enzyme annotated as dithiol–disulfide isomerase DsbA. It preserves a Cys–Pro–Phe–Cys active-site motif, which is presumed to be an exclusive characteristic of the novel DsbA–mycoredoxin 1 (Mrx1) cluster. However, the real mode of action, the nature of the electron donor pathway and biological functions of NCgl2478 in C. glutamicum have remained enigmatic so far. Herein, we report that NCgl2478 plays an important role in stress resistance. Deletion of the ncgl2478 gene increases the size of growth inhibition zones. The ncgl2478 expression is induced in the stress-responsive extra-cytoplasmic function-sigma (ECF-σ) factor SigH-dependent manner by stress. It receives electrons preferentially from the mycothiol (MSH)/mycothione reductase (Mtr)/NADPH pathway. Further, NCgl2478 reduces S-mycothiolated mixed disulfides and intramolecular disulfides via a monothiol–disulfide and a dithiol–disulfide exchange mechanism, respectively. NCgl2478 lacks oxidase activity; kinetic properties of its demycothiolation are different from those of Mrx1. Site-directed mutagenesis confirms Cys24 is the resolving Cys residue, while Cys21 is the nucleophilic cysteine that is oxidized to a sulfenic acid and then forms an intramolecular disulfide bond with Cys24 or a mixed disulfide with MSH under oxidative stress. In conclusion, our study presents the first evidence that NCgl2478 protects against various stresses by acting as an MSH-dependent thiol–disulfide reductase, belonging to a novel DsbA–Mrx1 cluster.

NAD(P)H-dependent thioredoxin-disulfide reductase TrxR is essential for tellurite and selenite reduction and resistance in Bacillus sp. Y3

ABSTRACT Microbial reduction of selenite [Se(IV)] and tellurite [Te(IV)] to Se(0) and Te(0) can function as a detoxification mechanism and serve in energy conservation. In this study, Bacillus sp. Y3 was isolated and demonstrated to have an ability of simultaneous reduction of Se(IV) and Te(IV) during aerobic cultivation, with reduction efficiencies of 100% and 90%, respectively. Proteomics analysis revealed that the putative thioredoxin disulfide reductase (TrxR) and sulfate and energy metabolic pathway proteins were significantly upregulated after the addition of Se(IV) and Te(IV). qRT-PCR also showed an increased trxR transcription level in the presence of Se(IV) and Te(IV). Compared with a wild-type Escherichia coli strain, the TrxR-overexpressed E. coli strain showed higher Se(IV) and Te(IV) resistance levels and reduction efficiencies. Additionally, the TrxR showed in vitro Se(IV) and Te(IV) reduction activities when NADPH or NADH were present. When NADPH was used as the electron donor, the optimum conditions for enzyme activities were pH 8.0 and 37°C. The Km values of Te(IV) and Se(IV) were 16.31 and 2.91 mM, and the Vmax values of Te(IV) and Se(IV) were 12.23 and 11.20 µM min−1 mg−1, respectively. The discovery of the new reductive enzyme TrxR enriches the repertoire of the bacterial Se(IV) and Te(IV) resistance and reduction mechanisms. Bacillus sp. Y3 can efficiently reduce Se(IV) and Te(IV) simultaneously. Strain Y3 provides potential applications for selenite and tellurite bioremediation. The TrxR enzyme shows high catalytic activity for reducing Se(IV) and Te(IV). The discovery of TrxR enriches the bacterial Se(IV) and Te(IV) reduction mechanisms.

Eclipta alba Derived Phytochemicals against Jaundice

Phytochemicals otherwise known as secondary metabolites means the small organic molecules that are not essential for growth, development and reproduction but can protect from various diseases. Infact these phytochemicals are the key source of medicine. Secondary metabolites from Eclipta alba plant extract are traditionally used to cure Jaundice which is caused by Leptospira interrogans. The curative property of Eclipta alba against Leptospira interrogans was proved by molecular docking method in “Biovia Discovery Studio”. “High positive values of -CDOCKER energy and -CDOCKER interaction energy” recommended that pentadecan, heptadecane, 6,10,14-trimethyl-2-petadecanone can successfully deactivate the thioredoxin-disulfide reductase enzyme thereby disrupting the cellular function as well as the lifecycle of causative organism.