Proteomic approach and expression analysis revealed the differential expression of predicted leptospiral proteases capable of ECM degradation

Dhandapani, Gunasekaran; Sikha, Thoduvayil; Pinto, Sneha M.; Kumar, Manish; Patel, Krishna; Kumar, Vikram; Tennyson, Jebasingh; Satheeshkumar, Padikara Kutty; Gowda, Harsha; Prasad, T. S. Keshava; Madanan, Madathiparambil G.

doi:10.1016/j.bbapap.2018.04.006

Cited by 8 publications

(7 citation statements)

References 62 publications

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“…[12] A wide array of molecules on the outer membrane (OM) of Leptospira are involved in functions that are essential for the survival of Leptospira. [13][14][15] Various outer membrane proteins (OMPs) are directly involved in bacterial pathogenesis via cell adhesion, proteolysis, and immune evasion. [16,17] Therefore, it is important to identify and characterize these molecules as potential vaccine candidates and diagnostic targets against leptospirosis.…”

Section: Introductionmentioning

confidence: 99%

Triton X‐114 Fractionated Subcellular Proteome of Leptospira interrogans Shows Selective Enrichment of Pathogenic and Outer Membrane Proteins in the Detergent Fraction

et al. 2020

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The Triton X-114-based solubilization and temperature-dependent phase separation of proteins is used for subcellular fractionation where, aqueous, detergent, and pellet fractions represents cytoplasmic, outer membrane (OM), and inner membrane proteins, respectively. Mass spectrometry-based proteomic analysis of Triton X-114 fractions of proteomic analysis of Leptospira interrogans identified 2957 unique proteins distributed across the fractions. The results are compared with bioinformatics predictions on their subcellular localization and pathogenic nature. Analysis of the distribution of proteins across the Triton X-114 fractions with the predicted characteristics is performed based on "number" of unique type of proteins, and "quantity" which represents the amount of unique protein. The highest number of predicted outer membrane proteins (OMPs) and pathogenic proteins are found in aqueous and pellet fractions, whereas detergent fraction representing the OM has the highest quantity of OMPs and pathogenic proteins though lower in number than the aqueous and pellet fractions. This leaves the possibility of an upsurge in pathogenic proteins and OMPs on the OM under pathogenic conditions suggesting their potential use to combat leptospirosis. Further, the Triton X-114 subcellular fractions are more correlated to enrichment of pathogenic proteins predicted by MP3 software than predicted localization.

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Section: Introductionmentioning

confidence: 99%

Triton X‐114 Fractionated Subcellular Proteome of Leptospira interrogans Shows Selective Enrichment of Pathogenic and Outer Membrane Proteins in the Detergent Fraction

et al. 2020

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“…Protein modifications indicate active proteins and their functional capabilities . Our previous results showed a quantitative difference in protein content across subcellular locations in Leptospira . ,, The active form of protein can be assessed with the type of modification, subcellular location, coexpressed molecules, interacting molecules, and the environment. Hence, identifying the PTM content of a protein at the subcellular level is more beneficial to finding the role and characteristics of the protein than the PTMs identified from the total protein extract.…”

Section: Discussionmentioning

confidence: 99%

Unique Posttranslational Modification Sites of Acetylation, Citrullination, Glutarylation, and Phosphorylation Are Found to Be Specific to the Proteins Partitioned in the Triton X-114 Fractions of Leptospira

et al. 2022

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Posttranslational modifications (PTMs) are decisive factors in the structure, function, and localization of proteins in prokaryotic and eukaryotic organisms. However, prokaryotic organisms lack subcellular organelles, and protein localization based on subcellular locations like cytoplasm, inner membrane, periplasm, and outer membrane can be accounted for functional characterization. We have identified 131 acetylated, 1182 citrullinated, 72 glutarylated, 5 palmitoylated, and 139 phosphorylated proteins from Triton X-114 fractionated proteins of Leptospira , the pathogen of re-emerging zoonotic disease leptospirosis. In total, 74.7% of proteins were found exclusively in different Triton X-114 fractions. Additionally, 21.9% of proteins in multiple fractions had one or more PTM specific to different Triton X-114 fractions. Altogether, 96.6% of proteins showed exclusiveness to different Triton X-114 fractions either due to the presence of the entire protein or with a specific PTM type or position. Further, the PTM distribution within Triton X-114 fractions showed higher acetylation in aqueous, glutarylation in detergent, phosphorylation in pellet, and citrullination in wash fractions representing cytoplasmic, outer membrane, inner membrane, and extracellular locations, respectively. Identification of PTMs in proteins with respect to the subcellular localization will help to characterize candidate proteins before developing novel drugs and vaccines rationally to combat leptospirosis.

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“…The experimental design for the simulation of the host physiological environment under in vitro conditions was performed with mid log phase culture of Leptospira as described previously with slight modifications . Mid log phase culture of Leptospira in EMJH medium supplemented with 1% BSA was incubated at 37 °C to induce physiological temperature (PT) , or added 150 mM NaCl to induce physiological osmolarity (PO) , or both together to induce physiological temperature and osmolarity (PTO). ,,, All were simultaneously incubated for 24 h with 4 biological replicates of 25 mL each.…”

Section: Methodsmentioning

confidence: 99%

“…Adler and his team sought to understand the alterations in proteins on the leptospiral outer membrane in response to physiologic temperatures, which showed that more OMPs, noncytoplasmic proteins, and lipoproteins were downregulated in response to the changes in temperature, which was attributed to immune-evasion tactics by the pathogen to strategically hide its surface proteins at the host’s physiological temperature . The upregulation of leptospiral immunoglobulin-like domain-containing proteins LigA and LigB in response to mimicked mammalian physiological osmolarity had been demonstrated at protein and mRNA levels. ,− Several genes were found to be DE under physiological osmolarity and temperature upshift, and it was hypothesized that these physiologic parameters work together synergistically to induce the expression of some pathogenic proteins, including proteases, which are essential for the successful development of infection. ,− …”

Section: Introductionmentioning

confidence: 99%

Physiological Temperature and Osmotic Changes Drive Dynamic Proteome Alterations in the Leptospiral Outer Membrane and Enhance Protein Export Systems

Phukan,

Sarma,

Rex

et al. 2023

J. Proteome Res.

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Leptospirosis, a remerging zoonosis, has no effective vaccine or an unambiguous early diagnostic reagent. Proteins differentially expressed (DE) under pathogenic conditions will be useful candidates for antileptospiral measures. We employed a multipronged approach comprising high-resolution TMT-labeled LC-MS/MS-based proteome analysis coupled with bioinformatics on leptospiral proteins following Triton X-114 subcellular fractionation of leptospires treated under physiological temperature and osmolarity that mimic infection. Although there were significant changes in the DE proteins at the level of the entire cell, there were notable changes in proteins at the subcellular level, particularly on the outer membrane (OM), that show the significance of subcellular proteome analysis. The detergent-enriched proteins, representing outer membrane proteins (OMPs), exhibited a dynamic nature and upregulation under various physiological conditions. It was found that pathogenic proteins showed a higher proportion of upregulation compared to the nonpathogenic proteins in the OM. Further analysis identified 17 virulent proteins exclusively upregulated in the outer membrane during infection that could be useful for vaccine and diagnostic targets. The DE proteins may aid in metabolic adaptation and are enriched in pathways related to signal transduction and antibiotic biosynthesis. Many upregulated proteins belong to protein export systems such as SEC translocase, T2SSs, and T1SSs, indicating their sequential participation in protein transport to the outer leaflet of the OM. Further studies on OM-localized proteins may shed light on the pathogenesis of leptospirosis and serve as the basis for effective countermeasures.

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Proteomic approach and expression analysis revealed the differential expression of predicted leptospiral proteases capable of ECM degradation

Cited by 8 publications

References 62 publications

Triton X‐114 Fractionated Subcellular Proteome of Leptospira interrogans Shows Selective Enrichment of Pathogenic and Outer Membrane Proteins in the Detergent Fraction

Triton X‐114 Fractionated Subcellular Proteome of Leptospira interrogans Shows Selective Enrichment of Pathogenic and Outer Membrane Proteins in the Detergent Fraction

Unique Posttranslational Modification Sites of Acetylation, Citrullination, Glutarylation, and Phosphorylation Are Found to Be Specific to the Proteins Partitioned in the Triton X-114 Fractions of Leptospira

Physiological Temperature and Osmotic Changes Drive Dynamic Proteome Alterations in the Leptospiral Outer Membrane and Enhance Protein Export Systems

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