Publications by authors named "Pramod Subedi"

6 Publications

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BcfH Is a Trimeric Thioredoxin-Like Bifunctional Enzyme with Both Thiol Oxidase and Disulfide Isomerase Activities.

Antioxid Redox Signal 2021 Apr 12. Epub 2021 Apr 12.

Department of Biochemistry and Genetics, La Trobe Institute for Molecular Science, La Trobe University, Bundoora, Australia.

Thioredoxin (TRX)-fold proteins are ubiquitous in nature. This redox scaffold has evolved to enable a variety of functions, including redox regulation, protein folding, and oxidative stress defense. In bacteria, the TRX-like disulfide bond (Dsb) family mediates the oxidative folding of multiple proteins required for fitness and pathogenic potential. Conventionally, Dsb proteins have specific redox functions with monomeric and dimeric Dsbs exclusively catalyzing thiol oxidation and disulfide isomerization, respectively. This contrasts with the eukaryotic disulfide forming machinery where the modular TRX protein disulfide isomerase (PDI) mediates thiol oxidation and disulfide reshuffling. In this study, we identified and structurally and biochemically characterized a novel Dsb-like protein from termed bovine colonization factor protein H (BcfH) and defined its role in virulence. In the conserved bovine colonization factor () fimbrial operon, the Dsb-like enzyme BcfH forms a trimeric structure, exceptionally uncommon among the large and evolutionary conserved TRX superfamily. This protein also displays very unusual catalytic redox centers, including an unwound α-helix holding the redox active site and a proline instead of the conserved -proline active site loop. Remarkably, BcfH displays both thiol oxidase and disulfide isomerase activities contributing to fimbrial biogenesis. Typically, oligomerization of bacterial Dsb proteins modulates their redox function, with monomeric and dimeric Dsbs mediating thiol oxidation and disulfide isomerization, respectively. This study demonstrates a further structural and functional malleability in the TRX-fold protein family. BcfH trimeric architecture and unconventional catalytic sites permit multiple redox functions emulating in bacteria the eukaryotic PDI dual oxidoreductase activity.
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http://dx.doi.org/10.1089/ars.2020.8218DOI Listing
April 2021

Impact of COVID-19 pandemic on socioeconomic and mental health aspects in Nepal.

Int J Soc Psychiatry 2020 12 10;66(8):748-755. Epub 2020 Jul 10.

Department of Biochemistry and Genetics, La Trobe Institute for Molecular Science, La Trobe University, Melbourne, VIC, Australia.

Background: Public health concern is increasing with recent rise in the number of COVID-19 cases in Nepal. To curb this pandemic, Nepal is facing some forms of lockdown, encouraging people to implement social distancing so as to reduce interactions between people which could eventually reduce the possibilities of new infection; however, it has affected the overall physical, mental, social and spiritual health of the people.

Methods: Published articles related to psychosocial effects due to COVID-19 and other outbreaks were searched and reviewed.

Conclusion: While many countries are supporting their citizens with sophisticated health safety-nets and various relief funds, some developing countries have unique challenges with vulnerable populations and limited resources to respond to the pandemic. This review presents the consequences of pandemic and lockdown on socioeconomic, mental health and other aspects in Nepalese society.
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http://dx.doi.org/10.1177/0020764020942247DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7443960PMC
December 2020

A Molecular Chameleon for Mapping Subcellular Polarity in an Unfolded Proteome Environment.

Angew Chem Int Ed Engl 2020 06 8;59(25):10129-10135. Epub 2020 Jan 8.

Department of Chemistry and Physics, La Trobe Institute for Molecular Science, La Trobe University, Melbourne, VIC, 3086, Australia.

Environmental polarity is an important factor that drives biomolecular interactions to regulate cell function. Herein, a general method of using the fluorogenic probe NTPAN-MI is reported to quantify the subcellular polarity change in response to protein unfolding. NTPAN-MI fluorescence is selectively activated upon labeling unfolded proteins with exposed thiols, thereby reporting on the extent of proteostasis. NTPAN-MI also reveals the collapse of the host proteome caused by influenza A virus infection. The emission profile of NTPAN-MI contains information of the local polarity of the unfolded proteome, which can be resolved through spectral phasor analysis. Under stress conditions that disrupt different checkpoints of protein quality control, distinct patterns of dielectric constant distribution in the cytoplasm can be observed. However, in the nucleus, the unfolded proteome was found to experience a more hydrophilic environment across all the stress conditions, indicating the central role of nucleus in the stress response process.
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http://dx.doi.org/10.1002/anie.201914263DOI Listing
June 2020

The Scs disulfide reductase system cooperates with the metallochaperone CueP in copper resistance.

J Biol Chem 2019 11 23;294(44):15876-15888. Epub 2019 Aug 23.

Department of Biochemistry and Genetics, La Trobe Institute for Molecular Science, La Trobe University, Kingsbury Drive, Bundoora, Victoria 3083, Australia

The human pathogen serovar Typhimurium ( Typhimurium) contains a complex disulfide bond (Dsb) catalytic machinery. This machinery encompasses multiple Dsb thiol-disulfide oxidoreductases that mediate oxidative protein folding and a less-characterized suppressor of copper sensitivity () gene cluster, associated with increased tolerance to copper. To better understand the function of the Scs system, here we characterized two of its key components, the membrane protein ScsB and the periplasmic protein ScsC. Our results revealed that these two proteins form a redox pair in which the electron transfer from the periplasmic domain of ScsB (n-ScsB) to ScsC is thermodynamically driven. We also demonstrate that the Scs reducing pathway remains separate from the Dsb oxidizing pathways and thereby avoids futile redox cycles. Additionally, we provide new insight into the molecular mechanism underlying Scs-mediated copper tolerance in We show that both ScsB and ScsC can bind toxic copper(I) with femtomolar affinities and transfer it to the periplasmic copper metallochaperone CueP. Our results indicate that the Scs machinery has evolved a dual mode of action, capable of transferring reducing power to the oxidizing periplasm and protecting against copper stress by cooperating with the regulon, a major copper resistance mechanism in Overall, these findings expand our understanding of the functional diversity of Dsb-like systems, ranging from those mediating oxidative folding of proteins required for infection to those contributing to defense mechanisms against oxidative stress and copper toxicity, critical traits for niche adaptation and survival.
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http://dx.doi.org/10.1074/jbc.RA119.010164DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6827279PMC
November 2019

Structural and biochemical insights into the disulfide reductase mechanism of DsbD, an essential enzyme for neisserial pathogens.

J Biol Chem 2018 10 4;293(43):16559-16571. Epub 2018 Sep 4.

From the Department of Biochemistry and Genetics, La Trobe Institute for Molecular Science, La Trobe University, Melbourne 3086, Victoria, Australia,

The worldwide incidence of neisserial infections, particularly gonococcal infections, is increasingly associated with antibiotic-resistant strains. In particular, extensively drug-resistant strains that are resistant to third-generation cephalosporins are a major public health concern. There is a pressing clinical need to identify new targets for the development of antibiotics effective against -specific processes. In this study, we report that the bacterial disulfide reductase DsbD is highly prevalent and conserved among spp. and that this enzyme is essential for survival of DsbD is a membrane-bound protein that consists of two periplasmic domains, n-DsbD and c-DsbD, which flank the transmembrane domain t-DsbD. In this work, we show that the two functionally essential periplasmic domains of DsbD catalyze electron transfer reactions through unidirectional interdomain interactions, from reduced c-DsbD to oxidized n-DsbD, and that this process is not dictated by their redox potentials. Structural characterization of the n- and c-DsbD domains in both redox states provides evidence that steric hindrance reduces interactions between the two periplasmic domains when n-DsbD is reduced, thereby preventing a futile redox cycle. Finally, we propose a conserved mechanism of electron transfer for DsbD and define the residues involved in domain-domain recognition. Inhibitors of the interaction of the two DsbD domains have the potential to be developed as anti-neisserial agents.
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http://dx.doi.org/10.1074/jbc.RA118.004847DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6204915PMC
October 2018

Autotransporter Adhesins in Escherichia coli Pathogenesis.

Proteomics 2017 Dec 12;17(23-24). Epub 2017 Oct 12.

Department of Biochemistry and Genetics, La Trobe Institute for Molecular Science, La Trobe University, Melbourne, Victoria, Australia.

Most bacteria produce adhesion molecules to facilitate the interaction with host cells and establish successful infections. An important group of bacterial adhesins belong to the autotransporter (AT) superfamily, the largest group of secreted and outer membrane proteins in Gram-negative bacteria. AT adhesins possess diverse functions that facilitate bacterial colonisation, survival and persistence, and as such are often associated with increased bacterial fitness and pathogenic potential. In this review, we will describe AIDA-I type AT adhesins, which comprise the biggest and most diverse group in the AT family. We will focus on Escherichia coli proteins and define general aspects of their biogenesis, distribution, structural properties and key roles in infection.
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http://dx.doi.org/10.1002/pmic.201600431DOI Listing
December 2017