Biology subjects

Dutta, D.

Publications and source records attributed to Dutta, D..

5 recordsLinked to original sources

bioRxiv2018

Comparative mode of action of antimicrobial peptide melimine and its derivative Mel4 against Pseudomonas aeruginosa

Melimine and Mel4 are chimeric cationic peptides with broad spectrum antimicrobial activity, and recent investigations have shown that they are highly biocompatible with animal model and human clinical trials. The current study examined the mechanism of action of these two antimicrobial peptides against P. aeruginosa with a series of investigations. Antimicrobial activities were determined by MIC and MBC. Endotoxin neutralization was determined using the LAL assay, effect on the cytoplasmic membrane was evaluated using DiSC(3)-5 and Sytox green stains, and Syto-9 and PI dyes using flow cytometry. Release of cytoplasmic materials (ATP and DNA/RNA) were determined using ATP luminescence and increase in OD260nm. The ability to lyse bacteria was studied by measuring a decrease in OD620nm. The MIC of the peptides remained low against P. aeruginosa strains, which showed efficient neutralization of LPS, indicating their role in the anti-pseudomonas and LPS binding activities. Both AMPs rapidly (starting at 30 seconds) depolarized P. aeruginosa cytoplasmic membrane leading to reduction in viability. Melimine was responsible for more ATP release (75%) compared to Mel4 (36%) (P<0.001) following two minutes exposure. For both peptides, Sytox green entered cells after five minutes of incubation. Flow cytometry demonstrated that both the AMPs permeabilized the cell membrane at 30 minutes and followed by increasing permeability. Similar results were found with DNA/RNA release experiments. Overall, melimine showed higher ability of membrane disruption, cell lysis compared to Mel4 (P<0.001). Knowledge regarding mechanism of action of these two AMPs would be helpful in making them as anti-pseudomonas drug.

microbiology↗

bioRxiv2018

Chemically Accurate Relative Folding Stability of RNA Hairpins from Molecular Simulations

This study describes a comparison between melts and simulated stabilities of the same RNAs that could be used to benchmark RNA force fields, and potentially to determine future melt-ing experiments. Using umbrella sampling molecular simulations of three 12-nucleotide RNA hairpin stem loops, for which there are experimentally determined free energies of unfold-ing, we projected unfolding onto the reaction coordinate of end to end (5' to 3' hydroxyl oxygen) distance. We estimate the free energy change of the transition from the native con-formation to a fully extended conformation--the stretched state--with no hydrogen bonds between non-neighboring bases. Each simulation was performed four times using the AM-BER FF99+bsc0+{chi}OL3 force field and each window, spaced at 1 [A] intervals, was sampled for 1 s, for a total of 552 s of simulation. We compared differences in the simulated free energy changes to analogous differences in free energies from optical melting experiments using ther-modynamic cycles where the free energy change between stretched and random coil sequences is assumed to be sequence independent. The differences between experimental and simulated {Delta}{Delta}G{degrees} are on average 1.00 {+/-} 0.66 kcal/mol, which is chemically accurate and suggests analo-gous simulations could be used predictively. We also report a novel method to identify where replica free energies diverge along the reaction coordinate, thus indicating where additional sampling would most improve convergence. We conclude by discussing methods to more economically perform such simulations.

biophysics↗

bioRxiv2018

SIV/SHIV-Zika coinfection does not alter disease pathogenesis in adult non-pregnant Rhesus Macaques

Due to the large geographical overlap of populations exposed to Zika virus (ZIKV) and human immunodeficiency virus (HIV), understanding disease pathogenesis in such coinfections is urgently needed. We used chronically infected simian immunodeficiency virus and chimeric simian human immunodeficiency virus (SIV/SHIV) macaques and inoculated with ZIKV. Plasma viral loads of both SIV/SHIV and ZIKV showed no significant changes as compared to ZIKV alone-infected animals. Tissue clearance of ZIKV was observed similarly. Furthermore, minimal changes in cytokines/chemokines were observed. Collectively, these data suggest that coinfection may not alter disease pathogenesis and warrants large HIV-ZIKV epidemiological studies to validate these findings.\n\nAuthor SummaryThe co-infection incidence of human immunodeficiency virus (HIV) infection and neglected tropical infectious diseases is increasing due to the large geographical overlap of populations exposed to both of these viruses. Thus, researching on such coinfection is of particular importance. In this study, we investigated HIV-ZIKV coinfection dynamics in adult non-pregnant Rhesus Macaques model chronically infected with simian immunodeficiency virus (SIV) - or chimeric simian human immunodeficiency virus (SHIV). We found that post ZIKV inoculation, plasma viral loads were similar to ZIKV alone infected animals in addition to minimal changes of cytokines. Dynamics of SIV and SHIV also did not change. Tissue clearance of ZIKV was found 67 months later. Our findings provide insights into HIV-ZKIV coinfection to determine the alteration of their pathogenesis.

microbiology↗

bioRxiv2018

Escherichia coli β-clamp slows down DNA polymerase I dependent nick translation while accelerating ligation

The nick translation property of DNA polymerase I (Pol I) ensures the maturation of Okazaki fragments by removing primer RNAs and facilitating ligation. However, prolonged nick translation traversing downstream DNA is an energy wasting futile process, as Pol I simultaneously polymerizes and depolymerizes at the nick sites utilizing energy-rich dNTPs. Using an in vitro assay system, we demonstrate that the {beta}-clamp of the Escherichia coli replisome strongly inhibits nick translation on the DNA substrate. To do so, {beta}-clamp inhibits the strand displacement activity of Pol I by interfering with the interaction between the finger subdomain of Pol I and the downstream primer-template junction. Conversely, {beta}-clamp stimulates the 5 exonuclease property of Pol I to cleave single nucleotides or shorter oligonucleotide flaps. This single nucleotide flap removal at high frequency increases the probability of ligation between the upstream and downstream DNA strands at an early phase, terminating nick translation. Besides {beta}-clamp-mediated ligation helps DNA ligase to seal the nick promptly during the maturation of Okazaki fragments.

biochemistry↗

bioRxiv2017

Multi-SKAT: General framework to test multiple phenotype associations of rare variants

In genetic association analysis, a joint test of multiple distinct phenotypes can increase power to identify sets of trait-associated variants within genes or regions of interest. Existing multi-phenotype tests for rare variants make specific assumptions about the patterns of association with underlying causal variants and the violation of these assumptions can reduce power to detect association. Here we develop a general framework for testing pleiotropic effects of rare variants on multiple continuous phenotypes using multivariate kernel regression (Multi-SKAT). Multi-SKAT models effect sizes of variants on the phenotypes through a kernel matrix and performs a variance component test of association. We show that many existing tests are equivalent to specific choices of kernel matrices with the Multi-SKAT framework. To increase power of detecting association across tests with different kernel matrices, we developed a fast and accurate approximation of the significance of the minimum observed p-value across tests. To account for related individuals, our framework uses random effects for the kinship matrix. Using simulated data and amino acid and exome-array data from the METSIM study, we show that Multi-SKAT can improve power over single-phenotype SKAT-O test and existing multiple phenotype tests, while maintaining type I error rate.\n\nGrant NumbersThis research was conducted with support from the following grants from National Institute of Health: O_LIR01 HG008773 (D.D. and S.L.)\nC_LIO_LIR01 LM012535 (D.D. and S.L.)\nC_LIO_LIR01 HG000376 (L.S. and M.B.)\nC_LIO_LIU01 DK062370 (L.S. and M.B.)\nC_LI

genetics↗