Person: TURAN, KADİR
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TURAN
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KADİR
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Publication Metadata only The Human MTCH2 Protein Has a Negative Effect On the Influenza a Virus Replication(2021-12-26) TURAN, KADİR; Ulupınar P., Çağlayan E., Turan K.Influenza A viruses have a segmented genome consists of eight single-stranded RNA molecules. Viral replication and transcription are carried out by viral RNA polymerase (RdRP), which consists of PB2, PB2 and PA subunits. During viral infection, RdRP and other viral proteins interact with several host proteins to perform their functions. In this work, mitochondrial carrier homolog 2 (MTCH2) protein, which was found to be associated with viral PA protein by yeast two-hybrid assay, was investigated its importance in terms of viral replication. In order to detect the effects of MTCH2 on virus replication in HeLa and HEK293 cells, the protein level was artificially changed. RNA interference and CRISPR/Cas9 techniques were used for down-regulation of the MTCH2. To increase the MTCH2, the HEK293 cells were transfected with pCHA-MTCH2 plasmid. The cells with altered MTCH2 transcript/protein level were infected with influenza A/WSN and A/DkPen viruses, and the viral replication was evaluated by detection of viral RNA/proteins with qPCR/Western blot techniques and plaque assays. In addition, the RdRP activities in these cells were determined by mini-replicon tests. A significant increase of viral mRNA/protein was observed in knockdown HeLa. It was shown that avian influenza A/DkPen replication was affected more. The over-expression of MTCH2 in HEK293 cells had a negative effect on the viral RdRP. These results showed that the MTCH2 is a negative cellular factor for influenza A virus. Although deletion of MTCH2 genes was detected in HEK293 cells knocked out by CRISPR/Cas9 technique, no negative effect on viral replication was observed. This situation is thought to be caused by heterozygosity or some other factors. From these results, it was concluded that the MTCH2 protein functions as a negative regulation factor for influenza A viruses. This work was supported by a grant of the Scientific and Technological Research Council of Turkey (SBAG-112S518).Keywords: Influenza a Viruses, MTCH2 Protein, Viral Rna Polymerase, PA ProteinPublication Open Access Heterologous expression and initial in silico characterization of a novel snakin-Z peptide(2023-08-01) TURAN, KADİR; Teker T., Albayrak G., Turan K.Heterologous expression of the plant-derived snakin-Z (SNK-Z) peptide was carried out inEscherichia coliBL21 and Mach 1 strains using the pET14b, and pGEX-6P-1 vectors, and its tertiary structure was determined. The most efficient production of the recombinant fusion peptide (GST/SNK-Z) in both strains was achieved by induction with 0.1 mM isopropyl β-D-1-thiogalactopyranoside at 32°C. GST/SNK-Z with 30.14kDa which yielded 6.5mg/L protein, was purified by affinity chromatography followed by dialysis. The GST tags were removed using PreScission protease. The IC50of GST/SNK-Z was calculated as 12.07 µM forStaphylococcus aureus(ATCC 25923) using nonlinear regression analysis. The secondary structure of recombinant SNK-Z consisted of α-helix and coil sites. It’s 3-D model was generated with a confidence score of -0.20 and a template modeling score of 0.69 ± 0.12. The predicted solvent accessibility and B-factor profile were detected.This is the first report of heterologous expression of SNK-Z inE. coli. Our study provides fundamental data for the large-scale production of SNK-Z, which has a high potential for use in the pharmaceutical industry because of its previously reported antimicrobial effects.Publication Metadata only TÜSEB destekli COVID-19 aşı projeleri(2021-09-09) TURAN, KADİR; Turan K.Publication Open Access Effects of some interferon-related proteins on influenza a viruse RNA polymerase activity(2022-01-01) TURAN, KADİR; Çağlayan E., TURAN K.© 2022, Turkish Pharmacists Association. All rights reserved.Objectives: Interferons (IFNs) are one of the most important components of innate immunity against viruses, especially those carrying the RNA genomes such as influenza viruses. Upon viral infection, the IFNs are rapidly secreted, inducing the expression of several genes in the target cells and establishing an antiviral state. In this study, the effects of proteins encoded by some IFN-related genes on influenza A virus RNA-dependent RNA polymerase enzyme were investigated. We evaluated the importance of these proteins in the pathogenesis of different influenza A virus types. Materials and Methods: The IFN-related genes were amplified by polymerase chain reaction from the HEK293 cDNA library and cloned into pCHA expression vector. The expression of genes and subcellular localizations of the proteins were determined by Western blotting and immunofluorescence staining, respectively. The effects of IFNs-related proteins on virus RdRP enzyme were determined by influenza A virus mini-replicons. Results: The study revealed that the influenza A virus infections significantly altered the transcript level of the IFN-related CCL5, IFIT1, IFIT3, IFITM3, and OAS1 genes in HEK293 cells. It was determined that the alteration of the gene expression was also related to the virus type. The mini-replicon assays showed that the transient expression of CCL5, IFI27, OAS1, IFITM3, IFIT1, and IFIT3 have inhibitory effects on WSN and/or DkPen type virus RdRP enzymes. We observed that the proteins except OAS1 inhibited WSN type RdRP enzyme at a higher level than that of DkPen enzyme. Conclusion: It was concluded that influenza A virus infection significantly alters the IFN-related gene expression in the cells. Most of the proteins encoded from these genes showed an inhibitory effect on the virus RdRP enzymes in the HEK293 cells. The inhibition of the influenza virus RdRP with IFN-related proteins may be the result of direct or indirect interactions between the host proteins and the viral enzyme subunits.