Biology subjects

Mukhopadhyay, B.

Publications and source records attributed to Mukhopadhyay, B..

5 recordsLinked to original sources

bioRxiv2026

Mechanistic insights into redox activity and catalytic determinants of the haloarchaeal flavin-dependent oxidoreductase HvFdR

Members of the FAD-dependent oxidoreductase family (IPR050260) play diverse and key roles in maintaining cellular redox balance, yet the functions of many distinct subgroups within this family remain unknown. Here, we define the biochemical and physiological functions of the Haloferax volcanii flavin-dependent oxidoreductase HvFdR (HVO_2345; fdr), a haloarchaeal member of a previously uncharacterized IPR050260 subgroup. HvFdR binds FAD and catalyzes NAD(P)H oxidase, diaphorase and ferredoxin reductase activities, with a kinetic preference for NADPH over NADH and catalytic properties that are strongly influenced by oxygen availability. Under stoichiometric conditions, HvFdR mediates reverse electron transfer to NADP, suggesting that intracellular nicotinamide nucleotide pools regulate electron flow bidirectionally. Consistent with this reversibility, HvFdR bound-FAD exhibits a low midpoint redox potential (-413 mV), supporting its capacity to function as an electron donor. Deletion of fdr impairs growth and elevates intracellular NADPH levels, consistent with a role for HvFdR in maintaining NADP(H) homeostasis. Conserved residues K47 and Y323 are identified as determinants of HvFdR electron transfer activity and may function as a regulatory gate that modulates electron flow while limiting excessive H2O2 production under aerobic conditions. Together, these findings establish HvFdR as an oxygen-responsive flavin-dependent oxidoreductase that contributes to cellular redox homeostasis and provides functional insight into a previously uncharacterized subgroup of the IPR050260 family.

biochemistry↗

bioRxiv2026

Lysine acetylation-mediated regulation of ferredoxin and ferredoxin reductase redox-active proteins in Haloferax volcanii

Lysine acetylation is an evolutionarily conserved, post-translational modification that regulates metabolism and protein function, yet its role in archaeal electron transfer systems remains poorly understood. Here, we investigated lysine acetylation of the 2Fe-2S ferredoxin HvFdx (HVO_2995) and its flavin-dependent oxidoreductase HvFdR (HVO_2345) partner in the halophilic archaeon Haloferax volcanii. Genetic and biochemical analyses established HvFdx as an essential 2Fe-2S ferredoxin with a midpoint redox potential of -385 mV. Lysine acetylation of HvFdx was found to occur primarily at K119, a residue positioned near the [Fe-S] cluster interface, and to modulate electron transfer capacity without impacting Fe-S cluster incorporation, midpoint potential, or protein abundance. In contrast, HvFdR was found lysine acetylated at multiple sites in a manner consistent with a non-enzymatic mechanism that resulted in altered flavin binding, enzymatic activity, and thermal stability. Lysine acetylation of HvFdx was found to stimulate electron flow from HvFdR as measured by an anaerobic NADPH [->] HvFdR [->] HvFdx [->] DCIP assay. 3D structural modeling, proteomic, biochemical, and genetic assays suggest the haloarchaeal GNAT-family acetyltransferase homolog HVO_2874 as a candidate enzyme associated with HvFdx lysine acetylation and optimal growth of H. volcanii. Together, these findings demonstrate that lysine acetylation differentially regulates archaeal redox-active proteins and functions as an important mechanism coordinating redox metabolism in H. volcanii.

microbiology↗

bioRxiv2020

Dominant Remodeling of Cattle Rumen Microbiome by Schedonorus arundinaceus (Tall Fescue) KY-31 Carrying a Fungal Endophyte

Tall fescue KY-31 feeds ~20% of the beef cattle in the United States. It carries a fungal endophyte that produces ergovaline, which causes toxicosis in cattle, leading to $2 billion revenue loss annually. The MaxQ cultivar of the grass is non-toxic, but less attractive economically. To develop ways of mitigating the toxicity, the rumen microbiome of cattle consuming KY-31 and MaxQ have been analyzed, principally for identifying ergovaline transforming microorganisms and often using fecal microbiome as a surrogate. We have hypothesized that KY-31 not only causes toxicosis, but also impacts rumen metabolism broadly, and tested the hypothesis by analyzing rumen microbiome compositions of cattle that grazed MaxQ with an intervening KY-31 grazing period with 16S rRNA-V4 element as identifier. We found that KY-31 remodeled the cellulolytic and saccharolytic communities substantially. This effect was not evident at whole microbiome levels but in the compositions of sessile and planktonic fractions. A move from MaxQ to KY-31 lowered the Firmicutes abundance in the sessile fraction and increased it in planktonic part and caused an opposite effect for Bacteroidetes, although the total abundances of these dominant rumen organisms remained unchanged. In the sessile fraction, the abundances of Fibrobacter, which degrades less degradable fibers, and certain cellulolytic Firmicutes such as Pseudobutyrivibrio and Butyrivibrio 2, dropped, and these losses were apparently compensated by increased occurrences of Eubacterium and specific Ruminococcaceae and Lachnospiraceae. In planktonic fraction the Tenericutes abundance increased as saccharolytic Bacteroidetes level dropped. Several potential ergovaline degraders were enriched. A return to MaxQ restored the original Firmicutes and Bacteroidetes distributions. However, the Fibrobacter and Butyrivibrio 2 abundances remained low and their substitutes maintained significant presence. The rumen microbiome was influenced minimally by animals fescue toxicosis and was distinct from previously reported fecal microbiomes in composition. In summary, KY-31 and MaxQ cultivars of tall fescue were digested in the cattle rumen with distinct consortia and the KY-31-specific features were dominant. The study highlighted the importance of analyzing sessile and planktonic fractions separately.

microbiology↗

bioRxiv2020

Catechol-Containing Compounds are a Broad Class of Amyloid Inhibitors: I. Redox State is a Key Determinant of the Activities of the Inhibitory Activities

Mechanisms of amyloid inhibition remains poorly understood, in part because most protein targets of amyloid assembly are partially unfolded or intrinsically disordered, which hinders detailed structural characterization of protein-inhibitor complexes and structural-based mechanistic elucidation. Herein we employed a small molecule screening approach to identify inhibitors against three prototype amyloidogenic proteins: amylin, A{beta} and tau. One remarkable class of inhibitors identified was catechol-containing compounds and redox-related quinones/anthraquinones. Further mechanistic studies determined that the redox state of the broad class of catechol-containing compounds is a key determinant of the amyloid inhibitor activities.

biochemistry↗

bioRxiv2019

Whole genome sequence analysis reveals broad distribution of the RtxA type 1 secretion system and four novel type 1 secretion systems throughout the Legionella genus

Type 1 secretion systems (T1SSs) are broadly distributed among bacteria and translocate effectors with diverse function across the bacterial cell membrane. Legionella pneumophila, the species most commonly associated with Legionellosis, encodes a T1SS at the lssXYZABD locus which is responsible for the secretion of the virulence factor RtxA. Many investigations have failed to detect lssD, the gene encoding the membrane fusion protein of the RtxA T1SS, in non-pneumophila Legionella, suggesting that this system is a conserved virulence factor in L. pneumophila. Here we discovered RtxA and its associated T1SS in a novel Legionella taurinensis strain, leading us to question whether this system may be more widespread than previously thought. Through a bioinformatic analysis of publicly available data, we classified and determined the distribution of four T1SSs including the RtxA T1SS and four novel T1SSs among diverse Legionella spp. The ABC transporter of the novel Legionella T1SS Legonella repeat protein secretion system (LRPSS) shares structural similarity to those of diverse T1SS families, including the alkaline protease T1SS in Pseudomonas aeruginosa. The Legionella bacteriocin (1-3) secretion systems (LB1SS-LB3SS) T1SSs are novel putative bacteriocin transporting T1SSs as their ABC transporters include C-39 peptidase domains in their N-terminal regions, with LB2SS and LB3SS likely constituting a nitrile hydratase leader peptide transport T1SSs. The LB1SS is more closely related to the colicin V T1SS in Escherichia coli. Of 45 Legionella spp. whole genomes examined, 19 (42%) were determined to possess lssB and lssD homologs. Of these 19, only 7 (37%) are known pathogens. There was no difference in the proportions of disease associated and non-disease associated species that possessed the RtxA T1SS (p = 0.4), contrary to the current consensus regarding the RtxA T1SS. These results draw into question the nature of RtxA and its T1SS as a genetic virulence determinant.

microbiology↗