Biology subjects

Lal, R.

Publications and source records attributed to Lal, R..

9 recordsLinked to original sources

bioRxiv2026

Machine Learning-Assisted Evolution of Broadly Functional Enzyme Libraries

Biocatalysis offers sustainable solutions to pressing challenges in chemical synthesis by exploiting the remarkable efficiency and selectivity of enzymes. Importantly, enzymes are able to accommodate non-native substrates and mediate transformations outside of their natural repertoire. Enzymes can be engineered for diverse applications by harnessing these promiscuous activities and optimizing them using directed evolution (DE). The success of a DE campaign, however, depends on the availability of a protein starting point that displays detectable levels of the desired function. To find a starting point, researchers often screen libraries of protein variants for novel activities, typically with low rates of success. Here, instead, we diversified the active site of a desirable parent protein and applied machine learning to generate informed, promiscuous libraries of protein variants. Specifically, we tested 26 different carbene and nitrene transfer reactions and used active learning-assisted directed evolution (ALDE) to generate optimized protoglobin variants with high activity across multiple reactions. We observed improvements in activity and selectivity for every reaction performed by the parent enzyme in at least one member of the ALDE-predicted libraries. Moreover, variants from these libraries can catalyze 5 out of 10 reactions not catalyzed by the parent protoglobin. These results indicate that supervised machine learning can help guide the construction of high-value enzyme libraries with expanded catalytic scope. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=99 SRC="FIGDIR/small/740427v1_ufig1.gif" ALT="Figure 1"> View larger version (31K): org.highwire.dtl.DTLVardef@fc2071org.highwire.dtl.DTLVardef@b5e787org.highwire.dtl.DTLVardef@737976org.highwire.dtl.DTLVardef@12b5e51_HPS_FORMAT_FIGEXP M_FIG C_FIG

bioengineering↗

bioRxiv2026

Strain-specific response to pharmacological cold mimicking in aging mice

BackgroundAging impairs thermogenic and metabolic flexibility, increasing metabolic disease risk. We examined six aging male mouse strains for responses to chronic topical menthol, a pharmacological cold mimetic. Material and methodsMale (C57BL/6J, A/J, BALB/c, C3H/hej, DBA/2J, and FVB/NJ) mice were treated with 4g/kg of 10% menthol weight per volume cream once per day or vehicle as control using finger application for 15 days. After last application, core body temperature measurement, BAT thermography, Nesting behaviour, and metabolic tolerance test were performed, and gene expression was analysed in BAT. ResultsMenthol improved core temperature recovery via thermogenesis and enhanced insulin sensitivity across all strains. However, effects on BAT mass, nesting behaviour, and gluconeogenesis were strain-dependent. ConclusionCollectively, these findings highlight that genetic background modulates metabolic responses to cold mimetics, informing future precision strategies for age-related metabolic disorders.

pharmacology and toxicology↗

bioRxiv2026

Role of brown adipose tissue-specific sympathetic and sensory innervations in menthol-induced energy-expending phenotype

Menthol, a TRPM8 agonist and pharmacological cold mimic, activates brown adipose tissue (BAT) to increase adaptive thermogenesis and whole-body energy expenditure. BAT is highly innervated with sensory and sympathetic nerve fibers. There is existing literature that sympathetic nerves have a critical role in regulating menthols (or cold) effect on energy homeostasis, but there is minimal literature about the involvement of somatosensory innervations. Here, we ought to investigate whether short-term topical application (3 days) of menthol elicits sympathetic and sensory nerves. We used chemical denervation models of complete (100 mg/kg; 6-OHDA and 125 mg/kg; capsaicin, s.c.) and localized (in BAT, 20 {micro}L of 6-OHDA (20 mg/ml) and 20{micro}L of capsaicin (20{micro}g/{micro}L) in each BAT lobe) ablation of sympathetic (6-OHDA) and sensory (capsaicin) innervations to explore their roles in menthol induced BAT activation and energy expanding phenotype. In the present study we have shown that (i) short-term topical application of menthol (10 % menthol for 3 consecutive days) elicits sympathetic and sensory innervations, and induces thermogenesis, lipolysis and mitochondrial biogenesis in mice BAT; (ii) localized ablation of sympathetic innervation prevented menthol induced BAT activation and lipolysis (iii) localized ablation of sensory neurons augments sympathetic innervations induced BAT activation and modulated thermoregulation. Collectively, our results suggest that BAT activation following short-term topical application of menthol is dependent primarily but not exclusively on centrally mediated SNS activation. TRPV1-positive sensory neurons are partially necessary for thermoregulation and efficient BAT activation.

pharmacology and toxicology↗

bioRxiv2023

Ketogenic diet prevents sucrose withdrawal-induced anxiety-like behaviour in male mice

Sugar bingeing has been shown to induce withdrawal effects when sugar is no longer available in the diet. In this study, we investigated the sex differences in sugar-addiction-like behaviour and the effects of the ketogenic diet on these outcomes. We used the two-bottle sucrose choice paradigm as a pre-clinical sucrose overeating and withdrawal model. Female mice consumed more sucrose than males when given free access to water and 10% sucrose for four weeks. One week of sucrose withdrawal after four weeks of consecutive sucrose overeating showed anxiety-like behaviour in male mice. However, the ketogenic diet did not affect sucrose overconsumption in males and females but prevented the sucrose withdrawal-induced anxiety-like behaviour in male mice with a concomitant increase in cFos mRNA in the prefrontal cortex. These findings provide evidence of sex differences in sucrose addiction-like behaviour and also indicate that a ketogenic diet may prevent sucrose withdrawal-induced anxiety-like behaviour in males. However, further research is needed to explore the underlying molecular mechanisms driving sex differences in sugar-addiction-like behaviour and the anxiolytic effects of the ketogenic diet.

neuroscience↗

bioRxiv2022

Monkey Pox Virus (MPXV): Phylogenomics, Host-Pathogen Interactome, and Mutational Cascade

While the world is still managing to recover from Covid-19 pandemic, Monkeypox awaits to bring in another global outbreak as a challenge to the entire mankind. However, Covid-19 pandemic have taught us lessons to move fast in viral genomic research to implement prevention and treatment strategies. One of the important aspects in Monkeypox virus should be immediately taken up is to gather insights of its evolutionary lineage based on the genomic studies. We have thus analysed the genome sequences of reported isolates of Monkeypox in the present study through phylogenomics. Host-pathogen interactions, mutation prevalence and evolutionary dynamics of this virus were investigated for all the documented isolates. Phylogenetic exploration revealed the clustering of strain Israel 2018 (MN 648051.1) from Clade I with the four isolates reported from the recent outbreak. An in-depth scrutiny of the host-pathogen interactome identified protein E3, serine protease inhibitor-2 (SPI-2), protein K7, and cytokine response-modifying protein B (CrmB) as the major regulatory hubs. Among these, the CrmB protein (dN/dS {approx} 1.61) was detected to be operating through positive selection. It possibly attests a selective advantage with the monkeypox virus in protecting the infected cells from antiviral responses elicited by the host. Studies also revealed that CrmB protein exhibited several mutations, the majority of which were destabilizing ({Delta}{Delta}G >0). While this study identified a large number of mutations within the newly outbreak clade, it also reflected that we need to move fast with the genomic analysis of the newly detected strains from around the world to develop better prevention and treatment methods

microbiology↗

bioRxiv2021

Microbial ecology of sulfur biogeochemical cycling at a mesothermic hot spring atop Northern Himalayas, India

O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=146 SRC="FIGDIR/small/470874v1_ufig1.gif" ALT="Figure 1"> View larger version (49K): org.highwire.dtl.DTLVardef@1dfd0d5org.highwire.dtl.DTLVardef@107811forg.highwire.dtl.DTLVardef@1ae4af6org.highwire.dtl.DTLVardef@1bb90a1_HPS_FORMAT_FIGEXP M_FIG Graphical Abstract C_FIG Sulfur Related Prokaryotes (SRP) residing in hot spring present good opportunity for exploring the limitless possibilities of integral ecosystem processes. Metagenomic analysis further expand the phylogenetic breadth of these extraordinary sulfur metabolizing microorganisms, as well a their complex metabolic networks and syntrophic interactions in environmental biosystems. Through this study, we explored and expanded the microbial genetic repertoire with focus on sulfur cycling genes through metagenomic analysis of sulfur (S) contaminated hot spring, located at the Northern Himalayas. The analysis revealed rich diversity of microbial consortia with established roles in S cycling such as Pseudomonas, Thioalkalivibrio, Desulfovibrio and Desulfobulbaceae (Proteobacteria). The major gene families inferred to be abundant across microbial mat, sediment and water were assigned to Proteobacteria as reflected from the RPKs (reads per kilobase) categorized into translation and ribosomal structure and biogenesis. Analysis of sequence similarity showed conserved pattern of both dsrAB genes (n=178) retrieved from all metagenomes while other sulfur disproportionation proteins were diverged due to different structural and chemical substrates. The diversity of sulfur oxidizing bacteria (SOB) and sulfate reducing bacteria (SRB) with conserved (r)dsrAB suggests for it to be an important adaptation for microbial fitness at this site. Here, we confirm that (i) SRBs belongs to{delta} -Proteobacteria occurring independent LGT of dsr genes to different and few novel lineages (ii) also, the oxidative and reductive dsr evolutionary time scale phylogeny, proved that the earliest (not first) dsrAB proteins belong to anaerobic Thiobacillus with other (rdsr) oxidizers. Further, the structural prediction of unassigned DsrAB proteins confirmed their relatedness with species of Desulfovibrio (TM score= 0.86; 0.98; 0.96) and Archaeoglobus fulgidus (TM score= 0.97; 0.98). We proposed that the genetic repertoire might provide the basis of studying time scale evolution and horizontal gene transfer of these genes in biogeochemical S cycling and the complementary genes could be implemented in biotechnology and bioremediation applications.

microbiology↗

bioRxiv2020

Multi-Omics and Integrated Network Approach to Unveil Evolutionary Patterns, Mutational Hotspots, Functional Crosstalk and Regulatory Interactions in SARS-CoV-2

SARS-CoV-2 pandemic resulted in 92 million cases in a span of one year. The study focuses on understanding population specific variations attributing its high rate of infections in specific geographical regions particularly in USA. Rigorous phylogenomic network analysis of complete SARS-CoV-2 genomes (245) inferred five central clades named a (ancestral), b, c, d and e (subtype e1 & e2). The clade d & e2 were found exclusively comprising of USA. Clades were distinguished by 10 co-mutational combinations in Nsp3, ORF8, Nsp13, S, Nsp12, Nsp2 and Nsp6. Our analysis revealed that only 67.46% of SNP mutations were at amino acid level. T1103P mutation in Nsp3 was predicted to increase protein stability in 238 strains except 6 strains which were marked as ancestral type; whereas co-mutation (P409L & Y446C) in Nsp13 were found in 64 genomes from USA highlighting its 100% co-occurrence. Docking highlighted mutation (D614G) caused reduction in binding of Spike proteins with ACE2, but it also showed better interaction with TMPRSS2 receptor contributing to high transmissibility among USA strains. We also found host proteins, MYO5A, MYO5B, MYO5C had maximum interaction with viral proteins (N, S, M). Thus, blocking the internalization pathway by inhibiting MYO5 proteins which could be an effective target for COVID-19 treatment. The functional annotations of the HPI network were found to be closely associated with hypoxia and thrombotic conditions confirming the vulnerability and severity of infection. We also screened CpG islands in Nsp1 & N conferring ability of SARS-CoV-2 to enter and trigger ZAP activity inside host cell. ImportanceIn the current study we presented a global view of mutational pattern observed in SARS-CoV-2 virus transmission. This provided a who-infect-whom geographical model since the early pandemic. This is hitherto the most comprehensive comparative genomics analysis of full-length genomes for co-mutations at different geographical regions specially in USA strains. Compositional structural biology results suggested that mutations have balance of contrary forces effect on pathogenicity suggesting only few mutations to effective at translation level but not all. Novel HPI analysis and CpG predictions elucidates the proof of concept of hypoxia and thrombotic conditions in several patients. Thus, the current study focuses the understanding of population specific variations attributing high rate of SARS-CoV-2 infections in specific geographical regions which may eventually be vital for the most severely affected countries and regions for sharp development of custom-made vindication strategies.

microbiology↗

bioRxiv2020

Comparative Genomic Analysis of Rapidly Evolving SARS CoV-2 Viruses Reveal Mosaic Pattern of Phylogeographical Distribution

The Coronavirus Disease-2019 (COVID-19) that started in Wuhan, China in December 2019 has spread worldwide emerging as a global pandemic. The severe respiratory pneumonia caused by the novel SARS-CoV-2 has so far claimed more than 60,000 lives and has impacted human lives worldwide. However, as the novel SARS-CoV-2 displays high transmission rates, their underlying genomic severity is required to be fully understood. We studied the complete genomes of 95 SARS-CoV-2 strains from different geographical regions worldwide to uncover the pattern of the spread of the virus. We show that there is no direct transmission pattern of the virus among neighboring countries suggesting that the outbreak is a result of travel of infected humans to different countries. We revealed unique single nucleotide polymorphisms (SNPs) in nsp13-16 (ORF1b polyprotein) and S-Protein within 10 viral isolates from the USA. These viral proteins are involved in RNA replication, indicating highly evolved viral strains circulating in the population of USA than other countries. Furthermore, we found an amino acid addition in nsp16 (mRNA cap-1 methyltransferase) of the USA isolate (MT188341) leading to shift in amino acid frame from position 2540 onwards. Through the construction of SARS-CoV-2-human interactome, we further revealed that multiple host proteins (PHB, PPP1CA, TGF-{beta}, SOCS3, STAT3, JAK1/2, SMAD3, BCL2, CAV1 & SPECC1) are manipulated by the viral proteins (nsp2, PL-PRO, N-protein, ORF7a, M-S-ORF3a complex, nsp7-nsp8-nsp9-RdRp complex) for mediating host immune evasion. Thus, the replicative machinery of SARS-CoV-2 is fast evolving to evade host challenges which need to be considered for developing effective treatment strategies.

microbiology↗

bioRxiv2019

Chemoinformatic-guided engineering of polyketide synthases

Polyketide synthase (PKS) engineering is an attractive method to generate new molecules such as commodity, fine and specialty chemicals. A significant challenge in PKS design is engineering a partially reductive module to produce a saturated {beta}-carbon through a reductive loop exchange. In this work, we sought to establish that chemoinformatics, a field traditionally used in drug discovery, could provide a viable strategy to reductive loop exchanges. We first introduced a set of donor reductive loops of diverse genetic origin and chemical substrate structures into the first extension module of the lipomycin PKS (LipPKS1). Product titers of these engineered unimodular PKSs correlated with atom pair chemical similarity between the substrate of the donor reductive loops and recipient LipPKS1, reaching a titer of 165 mg/L of short chain fatty acids produced by Streptomyces albus J1074 harboring these engineered PKSs. Expanding this method to larger intermediates requiring bimodular communication, we introduced reductive loops of divergent chemosimilarity into LipPKS2 and determined triketide lactone production. Collectively, we observed a statistically significant correlation between atom pair chemosimilarity and production, establishing a new chemoinformatic method that may aid in the engineering of PKSs to produce desired, unnatural products.

biochemistry↗