مقالات پذیرفته شده در ششمین کنگره بین المللی زیست پزشکی
Deciphering Therapeutic protein drugs via Environmental Proteomics research in marine invertebrates exposed to microbial contamination
Deciphering Therapeutic protein drugs via Environmental Proteomics research in marine invertebrates exposed to microbial contamination
Naghmeh Roohi-Shalmaee,1Rezvan Mousavi-Nadushan,2,*Pargol Ghavam Mostafavi,3Delavar Shahbazzadeh,4Kamran Pooshang Bagheri,5
1. Venom and Biotherapeutics Molecules Lab, Medical Biotechnology Department, Biotechnology Research Center, Pasteur Institute of Iran. Tehran, Iran. 2. Department of Marine Science, Faculty of Natural Resources and Environment, Tehran North Branch, Islamic Azad University Tehran, Iran. 3. Department of Marine Science, Faculty of Natural Resources and Environment, Science and Research Branch, Islamic Azad University, Tehran, Iran. 4. Venom and Biotherapeutics Molecules Lab., Biotechnology Dept., Biotechnology Research Center, Pasteur Institute of Iran. Tehran, Iran. 5. Venom and Biotherapeutics Molecules Lab., Biotechnology Dept., Biotechnology Research Center, Pasteur Institute of Iran. Tehran, Iran.
Introduction: The seas and oceans are the ultimate destination for pollutants, the costal sessile organisms have to acclimate or adapt to their conditions. Based on our field observations the Serpulids of the Persian Gulf has been constructed huge colonies in contaminated locations. In this study, we tried to use proteomics to understand the adaptation in this creature. So we first discovered the coelomic fluid of the polychaetes from polluted and non-polluted areas by (2DE), then followed by MALDI TOF analysis.
Methods: At first, we performed two dimensional electrophoresis. The coelomic fluid of the polychaetes of polluted and non-polluted areas, were conducted, and 2DE was performed in triplicate according to O’Farrell method. Then, the candidate spots were selected for further assay using MALDI-TOF analyses in an external facility (York University, UK). The candidate spots were cut from the 2-DE gel and digested by trypsin. The resultant peptide fragments were injected to a Bruker Ultrafex III-MALDI-TOF/TOF Mass Spectrometer (Bruker Co., USA). After that, we performed Bioinformatics analyses followed by (PMF) searching of databases and (BLAST) to provide protein annotation and approve concordance in protein identity. After Sequence homology searches, the evolutionary relationships of peptide fragments was controlled. To predict the precise functions of the proteins, we employed several servers for the accuracy of the work including ((CDD), PROSITE, InterPro, and panther servers.
Results: 2DE Image analysis revealed the highly abundant proteins and differentially expressed proteins in the specimens collected from the polluted area. So we found about Fifty-five spots from the samples of polluted area and 14 spots from the samples of non-polluted area. Four identical over expressed spots were selected as dominant biomarkers for further experiments and analyses. Then, MALDI TOF analysis of the peptide fragments derived from spot1, spot2, and spot3 showed homology with keratin (type II cytoskeletal), cyanelle 30S ribosomal protein S13, and peptidyl-prolyl cis–trans isomerase, respectively. Spot 4 showed no similarity with the sequences registered in NCBI and UniProt databases. BLASTP analyses confirmed that the sequence of peptide fragment derived from spot 1 had significant identity with keratin in other genera ranging from 100 to 75% (expectation value ranging from 2e−06 to 0.1), spot 2 had considerable identity with ‘30S ribosomal protein’ from 100 to 64% (E value ranging from 2e−06 to 0.2) and spot 3 had significant identity with ‘Peptidyl-prolyl cis–trans isomerase’ from 100 to 87% (E value ranging from 5e−10 to 1e−07). Also several servers ((CDD), PROSITE, InterPro, and panther servers confirmed the accuracy of family determination for identified sequences and the functionally important domains and sites.
Conclusion: The possible plastic adaptation against bacterial pollution (as an environmental stress), were investigated by 2-D electrophoresis. Spots with different expression were analyzed for Partial sequencing. The keratins may induce neutrophil degranulation and higher kinase activity in the cytoplasm of epidermal keratinocytes. Consequently, keratin may be considered as a new biomarker for detection of pollution stress and also as a probable sign of adaptation. 30S ribosomal protein S13 involve the binding of r-proteins to nucleic acids. Elevation of S13 indicates the elevation of protein or peptide translation referring to over-production of some peptides or proteins.Third spot, PPIases catalyzes the cis–trans isomerization of proline imidic peptide bonds in oligopeptides and accelerates folding of the produced peptides or proteins. It seems 3 identified peptides/proteins including keratin (type II cytoskeletal), cyanelle 30S ribosomal protein S13, and peptidyl-prolyl cis–trans isomerase are associated with the immune/defense system in humans and animals. These molecular responses may be referred to enhancing stress tolerance against bacterial pollution.