Search results for: prolyl oligopeptidase

Authors: Mohit Wadhawan, Sushma Rathaur

Abstract:

Lymphatic filariasis, also called elephantiasis is a tropical disease afflicting over 120 million people in 81 countries worldwide. Existing anti filarial drugs are effective against the larval stages of filarial parasites which call for an urgent need of drugs which are macrofilaricidal. Identification of molecular targets crucial for survival of filarial parasites is a prerequisite for drug designing. Prolyl oligopeptidase (POP) is one such crucial enzyme involved in the maturation and degradation of neuropeptides and peptide hormones. We have identified this peptidase in the bovine filarial parasite, Setaria cervi. Effect of inhibition of POP on the proteome profile of filarial parasite has been discussed in this study. Filarial parasites were exposed to Z-pro-prolinal (ZPP), a specific POP inhibitor for 8 h and the motility and viability of the parasites was observed. It significantly reduced the motility and viability of the parasites. To study the proteome profile, the cytosolic, endoplasmic reticulum (ER) and mitochondrial extracts of the adult female parasites were subjected to 2-dimensional electrophoresis. As analyzed by the PD-Quest software, the ZPP caused the alteration in the different subcellular proteins, and the significantly altered proteins were identified using MALDI-MS/MS spectrometry. The major proteins identified were found to play important role in diverse biological functions like signaling, redox regulation, energy metabolism, stress response, and cytoskeleton formation. Moreover, we found upregulation in the calcium binding proteins such as calreticulin, calponin, and calpain-6 suggesting that POP inhibition regulates calcium release. This relates to earlier reports that POP plays non-catalytic role in inositol 1,4,5-trisphosphate (IP3) signaling inducing release of calcium from ER. Taken together, the data demonstrated that inhibition of prolyl oligopeptidase alter the overall proteome signifying its role in survival of the filarial parasites. Thus this study provides a basis for the use of POP as a chemotherapeutic target for the treatment of lymphatic filariasis.

Keywords: lymphatic filariasis, setaria cervi, prolyl oligopeptidase, proteomics

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Authors: Vineeta Kaushik, Manisha Goel

Abstract:

Chaperones are proteins that help other proteins fold correctly, and are found in all domains of life i.e., prokaryotes, eukaryotes and archaea. Various comparative genomic studies have suggested that the archaeal protein folding machinery appears to be highly similar to that found in eukaryotes. In case of protein folding; slow rotation of peptide prolyl-imide bond is often the rate limiting step. Formation of the prolyl-imide bond during the folding of a protein requires the assistance of other proteins, termed as peptide prolyl cis-trans isomerases (PPIases). Cyclophilins constitute the class of peptide prolyl isomerases with a wide range of biological function like protein folding, signaling and chaperoning. Most of the cyclophilins exhibit PPIase enzymatic activity and play active role in substrate protein folding which classifies them as a category of molecular chaperones. Till date, there is not very much data available in the literature on archaeal cyclophilins. We aim to compare the structural and biochemical features of the cyclophilin protein from within the three domains to elucidate the features affecting their stability and enzyme activity. In the present study, we carry out in-silico analysis of the cyclophilin proteins to predict their conserved residues, sites under positive selection and compare these proteins to their bacterial and eukaryotic counterparts to predict functional divergence. We also aim to clone and express these proteins in heterologous system and study their biophysical characteristics in detail using techniques like CD and fluorescence spectroscopy. Overall we aim to understand the features contributing to the folding, stability and dynamics of the archaeal cyclophilin proteins.

Keywords: biophysical characterization, x-ray crystallography, chaperone-like activity, cyclophilin, PPIase activity

Procedia PDF Downloads 174

Authors: Jiang Xu, Daoping Wang, Yinghong Pan

Abstract:

The jointing-booting stage is a critical period of both vegetative growth and reproductive growth in rice. Therefore, the proteomic analysis of the mutant Osbri1, whose corresponding gene OsBRI1 encodes the putative BRs receptor OsBRI1, at jointing-booting stage is very important for understanding the effects of BRs on vegetative and reproductive growth. In this study, the proteomes of leaves from an allelic mutant of the DWARF 61 (D61, OsBRI1) gene, Fn189 (dwarf54, d54) and its wild-type variety T65 (Taichung 65) at jointing-booting stage were analysed by using a Q Exactive plus orbitrap mass spectrometer, and more than 3,100 proteins were identified in each sample. Ontology analysis showed that these proteins distribute in various space of the cells, such as the chloroplast, mitochondrion, and nucleus, they functioned as structural components and/or catalytic enzymes and involved in many physiological processes. Moreover, quantitative analysis displayed that 266 proteins were differentially expressed in two samples, among them, 77 proteins decreased and 189 increased more than two times in Fn189 compared with T65, the proteins whose content decreased in Fn189 including b5-like Heme/Steroid binding domain containing protein, putative retrotransposon protein, putative glutaminyl-tRNA synthetase, and higher content proteins such as mTERF, putative Oligopeptidase homologue, zinc knuckle protein, and so on. A former study founded that the transcription level of a mTERF was up-regulated in the leaves of maize seedling after EBR treatment. In our experiments, it was interesting that one mTERF protein increased, but another mTERF decreased in leaves of Fn189 at jointing-booting stage, which suggested that BRs may have differential regulation mechanisms on the expression of various mTERF proteins. The relationship between other differential proteins with BRs is still unclear, and the effects of BRs on rice protein contents and its regulation mechanisms still need further research.

Keywords: bri1 mutant, jointing-booting stage, proteomic analysis, rice

Procedia PDF Downloads 216

Authors: Umar Saeed

Abstract:

Peptidyl-prolyl cis/trans isomerases Par14 and Par17 in humans play crucial roles in diverse cellular processes, including protein folding, chromatin remodeling, DNA binding, ribosome biogenesis, and cell cycle progression. However, the effects of Par14 and Par17 on viral replication have been explored to a limited extent. We first time discovered their influential roles in promoting Hepatitis B Virus replication. In this study, we observed that in the presence of HBx, either Par14 or Par17 could upregulate HBV replication. However, in the absence of HBx, neither Par14 nor Par17 had any effect on replication. Their mechanism of action involves binding to specific motifs within HBc and HBx proteins. Notably, they target the conserved 133Arg-Pro134 (RP) motif of HBc and the 19RP20-28RP29 motifs of HBx. This interaction is fundamental for the stability of HBx, core particles, and HBc. Par14 and Par17 exhibit versatility by binding both outside and inside core particles, thereby facilitating core particle assembly through their participation in HBc dimer-dimer interactions. NAGE and immunoblotting analyses unveiled the binding of Par14/Par17 to core particles. Co-immunoprecipitation experiments further demonstrated the interaction of Par14/Par17 with core particle assembly-defective and dimer-positive HBc-Y132A. It's essential to emphasize that R133 is the key residue in the HBc RP motif that governs their interaction with Par14/Par17. Chromatin immunoprecipitation conducted on HBV-infected cells elucidated the participation of residues S19 and E46/D74 in Par14 and S44 and E71/D99 in Par17 in the recruitment of 133RP134 motif-containing HBc into cccDNA. Depleting PIN4 in liver cell lines results in a significant reduction in cccDNA levels, pgRNA, sgRNAs, HBc, core particle assembly, and HBV DNA synthesis. Notably, parvulin inhibitors like juglone and PiB have proven to be effective in substantially reducing HBV replication. These inhibitors weaken the interaction between HBV core particles and Par14/Par17, underscoring the dynamic nature of this interaction. It's also worth noting that specific Par14/Par17 inhibitors hold promise as potential therapeutic options for chronic hepatitis B.

Keywords: Par14Par17, HBx, HBc, cccDNA, HBV

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