Molecular Biology and Genetics / Moleküler Biyoloji ve Genetik
Permanent URI for this collectionhttps://hdl.handle.net/11147/9
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Article Citation - WoS: 9Citation - Scopus: 10Thiol Peroxidase Deficiency Leads To Increased Mutational Load and Decreased Fitness in Saccharomyces Cerevisiae(Genetics Society of America, 2014) Kaya, Alaattin; Lobanov, Alexey V.; Gerashchenko, Maxim V.; Koren, Amnon; Fomenko, Dmitri E.; Koç, Ahmet; Gladyshev, Vadim N.Thiol peroxidases are critical enzymes in the redox control of cellular processes that function by reducing low levels of hydroperoxides and regulating redox signaling. These proteins were also shown to regulate genome stability, but how their dysfunction affects the actual mutations in the genome is not known. Saccharomyces cerevisiae has eight thiol peroxidases of glutathione peroxidase and peroxiredoxin families, and the mutant lacking all these genes (Δ8) is viable. In this study, we employed two independent Δ8 isolates to analyze the genome-wide mutation spectrum that results from deficiency in these enzymes. Deletion of these genes was accompanied by a dramatic increase in point mutations, many of which clustered in close proximity and scattered throughout the genome, suggesting strong mutational bias. We further subjected multiple lines of wild-type and Δ8 cells to long-term mutation accumulation, followed by genome sequencing and phenotypic characterization. Δ8 lines showed a significant increase in nonrecurrent point mutations and indels. The original Δ8 cells exhibited reduced growth rate and decreased life span, which were further reduced in all Δ8 mutation accumulation lines. Although the mutation spectrum of the two independent isolates was different, similar patterns of gene expression were observed, suggesting the direct contribution of thiol peroxidases to the observed phenotypes. Expression of a single thiol peroxidase could partially restore the growth phenotype of Δ8 cells. This study shows how deficiency in nonessential, yet critical and conserved oxidoreductase function, leads to increased mutational load and decreased fitness.Article Citation - WoS: 5Citation - Scopus: 4Characterization of a Cdna From Beta Maritima That Confers Nickel Tolerance in Yeast(Elsevier Ltd., 2014) Bozdağ, Gönensin Ozan; Kaya, Alaattin; Koç, Ahmet; Noll, Gundula A.; Prüfer, Dirk; Karakaya, Hüseyin ÇağlarNickel is an essential micronutrient due to its involvement in many enzymatic reactions as a cofactor. However, excess of this element is toxic to biological systems. Here, we constructed a cDNA library from Beta maritima and screened it in the yeast system to identify genes that confer resistance to toxic levels of nickel. A cDNA clone (NIC6), which encodes for a putative membrane protein with unknown function, was found to help yeast cells to tolerate toxic levels of nickel. A GFP fused form of Nic6 protein was localized to multivesicular structures in tobacco epidermal cells. Thus, our results suggest a possible role of Nic6 in nickel and intracellular ion homeostasis.Article Citation - WoS: 11Citation - Scopus: 14Proteomic Changes During Boron Tolerance in Barley (hordeum Vulgare) and the Role of Vacuolar Proton-Translocating Atpase Subunit E(Türkiye Klinikleri Journal of Medical Sciences, 2011) Atik, Ahmet Emin; Bozdağ, Gönensin Ozan; Akıncı, Ersin; Kaya, Alaattin; Koç, Ahmet; Yalçın, Talat; Karakaya, Hüseyin ÇağlarBoron is an essential micronutrient for plants and animals; however, it can be toxic when present at high concentrations. The purpose of this study was to understand the mechanisms of boron tolerance in the Turkish barley (Hordeum vulgare) Anadolu cultivar. For this purpose, 2-dimensional electrophoresis (2-DE) was used to screen differentially expressed proteins for both control and boron-stressed Anadolu barley genotypes. Seven proteins were revealed by 2-DE: 1) ribulose 1,5-bisphosphate carboxylase/oxygenase (RuBisCo large chain), 2) TLP5, a thaumatin-like protein, 3) PR5, a basic pathogenesis-related protein, 4) a RNase S-like protein, 5) a PSI type III chlorophyll a/b-binding protein, 6) a light-harvesting complex I LHC I, and 7) the vacuolar proton-translocating ATPase subunit E protein. These were found to be upregulated in response to boron treatment. Even though the protein encoded by the V-ATPase subunit E gene was overexpressed, its transcript level was downregulated by boron treatment. Heterologous expression of the barley V-ATPase subunit E gene in yeast provided boron resistance to yeast cells. These results indicated that the V-ATPase subunit E gene was functional and conferred tolerance to toxic boron levels in yeast and might play a role in the overall boron tolerance of barley. © TÜBITAK.Article Citation - WoS: 73Citation - Scopus: 78Functional Analysis of Free Methionine-R Reductase From Saccharomyces Cerevisiae(American Society for Biochemistry and Molecular Biology, 2009) Le, Dung Tien; Lee, Byung Cheon; Marino, Stefano M.; Zhang, Yan; Fomenko, Dmitri E.; Kaya, Alaattin; Hacıoğlu, Elise; Kwak, Geun-Hee; Koç, Ahmet; Kim, Hwa-Young; Gladyshev, Vadim N.Methionine sulfoxide reductases (Msrs) are oxidoreductases that catalyze thiol-dependent reduction of oxidized methionines. MsrA and MsrB are the best known Msrs that repair methionine S-sulfoxide (Met-S-SO) and methionine-R-sulfoxide (Met-R-SO) residues in proteins, respectively. In addition, an Escherichia coli enzyme specific for free Met-R-SO, designated fRMsr, was recently discovered. In this work, we carried out comparative genomic and experimental analyses to examine occurrence, evolution, and function of fRMsr. This protein is present in single copies and two mutually exclusive subtypes in about half of prokaryotes and unicellular eukaryotes but is missing in higher plants and animals. A Saccharomyces cerevisiae fRMsr homolog was found to reduce free Met-R-SO but not free Met-S-SO or dabsyl-Met-R-SO. fRMsr was responsible for growth of yeast cells on Met-R-SO, and the double fRMsr/MsrA mutant could not grow on a mixture of methionine sulfoxides. However, in the presence of methionine, even the triple fRMsr/MsrA/MsrB mutant was viable. In addition, fRMsr deletion strain showed an increased sensitivity to oxidative stress and a decreased life span, whereas overexpression of fRMsr conferred higher resistance to oxidants. Molecular modeling and cysteine residue targeting by thioredoxin pointed to Cys101 as catalytic and Cys125 as resolving residues in yeast fRMsr. These residues as well as a third Cys, resolving Cys91, clustered in the structure, and each was required for the catalytic activity of the enzyme. The data show that fRMsr is the main enzyme responsible for the reduction of free Met-R-SO in S. cerevisiae.