Biology subjects

Mohan, S. V.

Publications and source records attributed to Mohan, S. V..

4 recordsLinked to original sources

bioRxiv2025

A Highly Thermostable and Novel GH5 Endoglucanase from Bacillus sp. Strain BS with Enhanced Biomass Saccharification Potential in Seawater

The efficient enzymatic conversion of lignocellulosic biomass to glucose is crucial for biofuel production, with enzyme costs being a significant barrier to commercialization. Multifunctional enzymes that reduce enzyme components can improve efficiency. In this study, we amplified and sequenced a gene from Bacillus sp. strain BS, which showed 89.8% identity with bacterial endoglucanase Q6YK34. This putative novel endoglucanase, BsEG2, is a GH5-like endoglucanase, with optimal activity at pH 6.0 and 55 {square}, and efficiently hydrolyzes amorphous and crystalline cellulose, including pre-treated and untreated sugarcane bagasse. Kinetics on carboxymethyl cellulose (CMC) indicate high substrate binding efficiency (apparent Vmax of 148.9 {micro}M min-1, a Km of 24.9 mg mL-1, and a kcat of 517 s-1). BsEG2 hydrolyzes long cellulosic chains, oligosaccharides, and cellobiose to produce glucose, demonstrating multifunctionality. It exhibits remarkable thermal stability, retaining over 90% activity after 15 days at 55 {degrees}C. BsEG2 also retains activity in high salts and ionic liquids, and is not inhibited by cellobiose, even at 200 mM, unlike other endoglucanases and cellobiohydrolases. Furthermore, BsEG2 functions in seawater, reducing freshwater footprint in biofuel production. BsEG2 is a promising candidate for a cost-effective enzyme formulation for glucose production from lignocellulosic biomass, supporting downstream glucose-fed metabolic pathways.

biochemistry↗

bioRxiv2024

Heterologous Expression of the carbon monoxide dehydrogenase Gene from Clostridium sp. to Enhance Acetic Acid and Alcohol Production from CO2

This study evaluates the performance of carbon monoxide dehydrogenase (codh) embedded strains in bench-scale microbial electrochemical systems (MES) for CO2 reduction to biofuels and biochemicals. CO2 fermentation efficiency was evaluated by comparing the wild-type Clostridium acetobutylicum (Wild), a negative control E. coli strain lacking the codh gene (NC-BL21), and engineered E. coli strain (Eng) alone and with IPTG induction (Eng+IPTG). Four electrochemical systems were used viz. Wild+E, NC-BL21+E, Eng+E, and Eng+IPTG+E, with a poised potential of 0.6 V applied to the working electrode. CO2 and bicarbonate were supplemented to a total inorganic carbon (IC) concentration of 40 g/L, with a retention time of 60 h. The engineered strains demonstrated enhanced metabolic performance compared to the wild-type and negative control strains, yielding a maximum formic acid concentration of 2.1 g/L and acetic acid concentration of 7.8 g/L under the Eng+IPTG condition. Ethanol yield was highest at 3.9 g/L under the Eng+IPTG+E condition, substantially exceeding the 2.4 g/L acetic acid yield observed in the wild-type strain. The engineered strains showed superior cumulative yields (0.4075 g/g), improved codh charge flux stability (60 vs. 5 for Wild), and upregulated expression of genes in the Wood-Ljungdahl pathway. Bioelectrochemical performance analysis demonstrated elevated reductive catalytic currents, enhanced CO2 reduction, and optimal charge transfer kinetics. This study highlights the effectiveness of genetic and process engineering, particularly codh overexpression and IPTG induction, in optimizing microbial electrosynthesis for biofuel and biochemical production from C1 gases.

bioengineering↗

bioRxiv2022

Codon optimality is the primary contributor to the exceptional mutational sensitivity of CcdA antitoxin in its operonic context

Deep mutational scanning studies suggest that synonymous mutations are typically silent and that most exposed, non active-site residues are tolerant to mutations. Here we show that the ccdA antitoxin component of the E.coli ccdAB toxin-antitoxin system is unusually sensitive to mutations when studied in the operonic context. A large fraction ([~]80%) of single-codon mutations, including many synonymous mutations in the ccdA gene shows inactive phenotype, but they retain native-like binding affinity towards cognate toxin, CcdB. Therefore, the observed phenotypic effects are largely not due to alterations in protein structure/stability, consistent with a large region of CcdA being intrinsically disordered. E. coli codon preference and strength of ribosome-binding associated with translation of downstream ccdB gene are found to be major contributors of the observed mutant phenotypes. In select cases, proteomics studies reveal altered ratios of CcdA:CcdB protein levels in vivo, suggesting that the ccdA mutations likely alter relative translation efficiencies of the two genes in the operon. We extend these results by studying single-site synonymous mutations that lead to loss of function phenotypes in the relBE operon upon introduction of rarer codons. Thus, in their operonic context, genes are likely to be more sensitive to both synonymous and non-synonymous point mutations than inferred previously.

molecular biology↗

bioRxiv2019

Loss of function from widely distributed, synonymous mutations at single codons

Mutational tolerance inferred from laboratory-based mutational studies is typically much higher than observed natural sequence variation. Using saturation mutagenesis, we show that the ccdA antitoxin component of the ccdAB toxin-antitoxin system is unusually sensitive to mutation with over 60% of mutations leading to loss of function. Multi-base synonymous mutations at a codon display enhanced propensity to show altered phenotypes, relative to single-base ones. Such mutations modulate RNA structure, leading to altered relative translation efficiencies of the two genes in the operon, and a CcdA:CcdB protein ratio below one. These insights were used to predict and experimentally validate synonymous mutations that lead to loss of function in the unrelated relBE operon as well as the lacZ gene. Thus, synonymous mutations can have significant phenotypic effects, in the absence of overexpression or extraneous reporters. More generally, proteins are likely more sensitive to mutation than inferred from previous saturation mutagenesis studies.

molecular biology↗