Biology subjects

Lynn, A. M.

Publications and source records attributed to Lynn, A. M..

5 recordsLinked to original sources

bioRxiv2022

Genome-wide analysis of PTR/POT transporters in Candida species and their functional characterization in the newly emerged pathogen Candida auris

The PTR or proton-dependent oligopeptide transporter (POT) family exploits the inwardly directed proton motive force to facilitate the cellular uptake of di- and tripeptides. Interestingly, representatives from the family have shown efficacy in delivering peptide-based antifungal derivatives in certain Candida species. Given the increased incidences of fungal infections by Candida species and the associated escalating orders of resistance against frontline antifungals, peptide derivatives are attractive therapeutic options. In that direction, the identification and characterization of PTR transporters serve as an essential first step in the translation of peptide-based antifungals as next-gen therapeutics. Herein, we present a genome-wide inventory of the PTR transporters in five prominent Candida species. Our study identifies 2 PTR transporters each in C. albicans and C. dubliniensis, 1 in C. glabrata, 4 in C. parapsilosis, and 3 in C. auris. Notably, despite all representatives retaining the conserved features seen in the PTR family, there exist two distinct classes of PTR transporters that differ in terms of their sequence identities and membrane topology. Further, we also evaluated the contribution of each PTR protein of the newly emerged multidrug-resistant C. auris in di-/tripeptide uptake. Notably, deletion of the PTR transporters encoded by BNJ08_003830 and BNJ08_005124 led to a marked reduction in the transport capabilities of several tested di-/tripeptides. Besides, BNJ08_005124 deletion also resulted in increased resistance towards the peptide-nucleoside drug Nikkomycin Z, pointing towards its predominant role in the uptake mechanism. Altogether, the study provides an important template for future structure-function investigations of PTR transporters in Candida species.

microbiology↗

bioRxiv2022

Genomic landscape of the DHA1 family in the newly emerged pathogen Candida auris and mapping substrate repertoire of the prominent member CauMdr1

The last decade has witnessed the rise of extremely threatening healthcare-associated multidrug resistant non-albicans Candida (NAC) species, Candida auris. Thus, understanding the molecular basis of antifungal resistance has emerged as the single most important goal amongst the research community. Since besides target alterations, efflux mechanisms contribute maximally to antifungal resistance, it is imperative to investigate their contributions in this pathogen. Of note, within the Major Facilitator Superfamily (MFS) of efflux pumps, Drug/H+ antiporter family 1 (DHA1) has been established as a predominant contributor towards xenobiotic efflux. Our study provides a complete landscape of DHA1 transporters encoded in the genome of C. auris. This study identifies 14 DHA1 transporters encoded in the genome of the pathogen. We also construct deletion and heterologous overexpression strains for the most important DHA1 drug transporter, viz., CauMdr1 to map the spectrum of its substrates. While the knockout strain did not show any significant changes in the resistance patterns against the majority of the tested substrates, the ortholog when overexpressed in a minimal background S. cerevisiae strain, AD1-8u-, showed significant enhancement in the Minimum inhibitory concentrations (MICs) against a large panel of antifungal molecules. Altogether, the present study provides a comprehensive template for investigating the role of DHA1 members of C. auris in antifungal resistance mechanisms.

microbiology↗

bioRxiv2020

Standardizing a protocol for studying pH-dependent transient conformations using computer simulations

1.Understanding pH-dependent protein stability is important in biological - transport, storage, and delivery, in vivo conditions such as aggregation potential in neurodegenerative disease, and in studying the folding/unfolding of proteins. Using computer simulations, we can replace complex experimental determination and provide an atomistic-level interpretation of the cause and effect of pH on protein stability. Here, we standardize a method that provides a framework through which we examined pH-dependent transient conformations during unfolding simulations of proteins. Constant pH simulations utilized in the prediction of pKa values of charged groups of the peptide. The calculated pKa values employed to fix the appropriate protonation state of the amino acid to simulate the effect of pH on the system. Trajectories from multiple high-temperature MD simulations of the protein sample the conformational space during unfolding for a given pH state. The ensemble of conformations is analyzed from its free energy landscape to identify transient and stable conformations both at a given pH and between different pH. As a test system RN80, a protein fragment analog of the C-peptide from bovine pancreatic ribonuclease-A used to measure the accuracy of the predictions from simulations. Experimental measures of the helix content determined as a function of pH display a bell-shaped curve, i.e. RN80 alpha-helix formation is maximum at pH5 with a subsequent loss in helicity at higher and lower pH. The main forces stabilizing the alpha-helix are a salt-bridge formed between Glu-2 and Arg-10 and cation-pi-interaction between Tyr-8 and His-12. Our protocol includes constant pH calculations, optimal high-temperature simulations, and Free Energy landscape analysis exhibited the agreement with the experimental observations.

bioinformatics↗

bioRxiv2020

ABC-finder: A containerized web server for the identification and topology prediction of ABC proteins

In view of the multiple clinical and physiological implications of ABC transporter proteins, there is a considerable interest among researchers to characterize them functionally. However, such characterizations are based on the premise that ABC proteins are accurately identified in the proteome of an organism, and their topology is correctly predicted. With this objective, we have developed ABC-finder, i.e., a Docker-based package for the identification of ABC proteins in all organisms, and visualization of the topology of ABC proteins using a web browser. ABC-finder is built and deployed in a Linux container, making it scalable for many concurrent users on our servers and enabling users to download and run it locally. Overall, ABC-finder is a convenient, portable, and platform-independent tool for the identification and topology prediction of ABC proteins. ABC-finder is accessible at http://abc-finder.osdd.jnu.ac.in

bioinformatics↗

bioRxiv2020

Modulation of macrophage defense responses by Mycobacterial persistence protein MprA (Rv0981) in human THP-1 cells: effect of single amino acid variation on host-pathogen interactions

M. tuberculosis is one of the most successful human pathogens causing tuberculosis that leads to highest daily morbidity worldwide. The evasion of the host immune responses is an important strategy that M. tuberculosis adopts. MprA (Rv0981), the response regulator of two component system is known for DNA binding activity in the pathogen and its role in persistent infection in the host. MprA is recognized as a late stage antigen during infection. A variant form of the protein MprA with G70S polymorphism (MprA*) is observed in one of our local and in several global clinical isolates of M. tuberculosis. Here we report the nuclear localization of MprA and MprA* in differentiated macrophages. MprA and MprA* increase the expression of TGF-{beta} and IL-10, the immune suppressive cytokines in THP-1 derived macrophage cells. Concurrently the phago-lysosome fusion is significantly reduced as shown by infection with M.bovis BCG. We show that single nucleotide variation in clinical isolates lead to quantitative variations resulting in host immune suppression and support the survival and persistence of the pathogen.

microbiology↗