Researcher:
Seferoğlu, Ayşe Bengisu

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PhD Student

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Ayşe Bengisu

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Seferoğlu

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Seferoğlu, Ayşe Bengisu

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2012 - 20175

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Researcher Of Publications: search.filters.isAuthorOfPublication.8c85e154-0885-4397-8095-7988d1db958b

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    Glu-370 in the large subunit influences the substrate binding, allosteric, and heat stability properties of potato ADP-glucose pyrophosphorylase
    (Elsevier Ireland Ltd, 2016) Çalışkan, Mahmut; Cevahir, Gül; N/A; Department of Chemical and Biological Engineering; N/A; Department of Molecular Biology and Genetics; N/A; Department of Chemical and Biological Engineering; Seferoğlu, Ayşe Bengisu; Gül, Şeref; Dikbaş, Uğur Meriç; Barış, İbrahim; Koper, Kaan; Kavaklı, İbrahim Halil; PhD Student; Researcher; Master Student; Teaching Faculty; Master Student; Faculty Member; Department of Molecular Biology and Genetics; Department of Chemical and Biological Engineering; Graduate School of Sciences and Engineering; College of Engineering; Graduate School of Sciences and Engineering; College of Sciences; Graduate School of Sciences and Engineering; College of Engineering; N/A; 289253; N/A; 111629; N/A; 40319
    ADP-glucose pyrophosphorylase (AGPase) is a key allosteric enzyme in plant starch biosynthesis. Plant AGPase is a heterotetrameric enzyme that consists of large (LS) and small subunits (SS), which are encoded by two different genes. In this study, we showed that the conversion of Glu to Gly at position 370 in the LS of AGPase alters the heterotetrameric stability along with the binding properties of substrate and effectors of the enzyme. Kinetic analyses revealed that the affinity of the (LSSSWT)-S-E370G AGPase for glucose 1-phosphate is 3-fold less than for wild type (WT) AGPase. Additionally, the (LSSSWT)-S-E370G AGPase requires 3-fold more 3-phosphogyceric acid to be activated. Finally, the LS(E370G)SS(WT)AGPase is less heat stable compared with the WT AGPase. Computational analysis of the mutant Gly-370 in the 3D modeled LS AGPase showed that this residue changes charge distribution of the surface and thus affect stability of the LS AGPase and overall heat stability of the heterotetrameric AGPase. In summary, our results show that LSE370 intricately modulate the heat stability and enzymatic activity of potato the AGPase.
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    Transcriptional regulation of the ADP-glucose pyrophosphorylase isoforms in the leaf and the stem under long and short photoperiod in lentil
    (Elsevier Ireland Ltd, 2013) Morgil, Hande; Tulum, Işıl; Cevahir, Gül; N/A; Department of Molecular Biology and Genetics; Department of Chemical and Biological Engineering; Seferoğlu, Ayşe Bengisu; Barış, İbrahim; Özdaş, Şule Beyhan; Kavaklı, İbrahim Halil; PhD Student; Teaching Faculty; Researcher; Faculty Member; Department of Molecular Biology and Genetics; Department of Chemical and Biological Engineering; Graduate School of Sciences and Engineering; College of Sciences; Graduate School of Sciences and Engineering; College of Engineering; N/A; 111629; 40319
    ADP-glucose pyrophosphorylase (AGPase) is a key enzyme in plant starch biosynthesis. It contains large (LS) and small (SS) subunits encoded by two different genes. In this study, we explored the transcriptional regulation of both the LS and SS subunits of AGPase in stem and leaf under different photoperiods length in lentil. To this end, we first isolated and characterized different isoforms of the LS and SS of lentil AGPase and then we performed quantitative real time PCR (qPCR) to see the effect of photoperiod length on the transcription of the AGPase isforms under the different photoperiod regimes in lentil. Analysis of the qPCR results revealed that the transcription of different isoforms of the LSs and the SSs of lentil AGPase are differentially regulated when photoperiod shifted from long-day to short-day in stem and leaves. While transcript levels of LS1 and SS2 in leaf significantly decreased, overall transcript levels of SS1 increased in short-day regime. Our results indicated that day length affects the transcription of lentil AGPase isoforms differentially in stems and leaves most likely to supply carbon from the stem to other tissues to regulate carbon metabolism under short-day conditions.
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    Identification of important residues for modulation of allosteric properties of potato ADP glucose pyrophosphorylase
    (Elsevier Science Bv, 2012) N/A; Department of Molecular Biology and Genetics; Department of Chemical and Biological Engineering; Seferoğlu, Ayşe Bengisu; Barış, İbrahim; Kavaklı, İbrahim Halil; PhD Student; Teaching Faculty; Faculty Member; Department of Molecular Biology and Genetics; Department of Chemical and Biological Engineering; Graduate School of Sciences and Engineering; College of Sciences; College of Engineering; N/A; 111629; 40319
    ADP glucose pyrophosphorylase (AGPase) is a key regulatory enzyme of bacterial glycogen and plant starch synthesis as it controls carbon flux via its allosteric regulatory behavior. Whereas the bacterial enzyme is composed of a single subunit type, the plant AGPase is a heterotetrameric enzyme (α2β2) with distinct roles for each of the two subunit types. The large subunit (LS) is involved mainly in allosteric regulation through its interaction with the catalytic small subunit (SS). Previously, critical amino acids of potato (Solanum tuberosum L.) LS that interact with SS in the native heterotetramer structure were identified both computationally and experimentally. In this study, we aimed to improve the heterotetrameric assembly of potato AGPase and to detect residues located on the interface involving the allosteric regulation of the enzyme with a reverse genetics approach. A mutant, α2β2 formation deficient, large subunit of potato AGPase named LSR88A was subjected to random mutagenesis using error prone PCR and screened for the capacity to form an enzyme restoring glycogen production in glgC-Escherichia coli, AGPase activity deficient, containing wild type SS by assessing iodine staining. Fifteen suppressor mutants were identified and sequence analysis of these mutants revealed that mutations are mainly clustered at subunit interface and nearby the subunit interface. Our kinetic characterization results indicate that interfaces between the large and small subunits are significant for the allosteric properties of the AGPase. Obtaining stable and up-regulated AGPase variants will enable us to use these mutants to increase the starch yield in crop plants.
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    Enhanced heterotetrameric assembly of potato ADP-Glucose pyrophosphorylase using reverse genetics
    (Oxford Univ Press, 2014) Cevahir, Gül; N/A; N/A; Department of Molecular Biology and Genetics; Department of Chemical and Biological Engineering; Seferoğlu, Ayşe Bengisu; Koper, Kaan; Can, Fatma Betül; Kavaklı, İbrahim Halil; PhD Student; Master Student; Undergraduate Student; Faculty Member; Department of Molecular Biology and Genetics; Department of Chemical and Biological Engineering; Graduate School of Sciences and Engineering; Graduate School of Sciences and Engineering; College of Sciences; College of Engineering; N/A; N/A; N/A; 40319
    ADP-glucose pyrophosphorylase (AGPase) is a key allosteric enzyme in plant starch biosynthesis. Plant AGPase is a heterotetrameric enzyme that consists of large (LS) and small subunits (SS), which are encoded by two different genes. Computational and experimental studies have revealed that the heterotetrameric assembly of AGPase is thermodynamically weak. Modeling studies followed by the mutagenesis of the LS of the potato AGPase identified a heterotetramer-deficient mutant, LSR88A. To enhance heterotetrameric assembly, LSR88A cDNA was subjected to error-prone PCR, and second-site revertants were identified according to their ability to restore glycogen accumulation, as assessed with iodine staining. Selected mutations were introduced into the wild-type (WT) LS and co-expressed with the WT SS in Escherichia coli glgC(-). The biochemical characterization of revertants revealed that (LSSSWT)-S-I90V, (LSSSWT)-S-Y378C and (LSSSWT)-S-D410G mutants displayed enhanced heterotetrameric assembly with the WT SS. Among these mutants, (LSSSWT)-S-Y378C AGPase displayed increased heat stability compared with the WT enzyme. Kinetic characterization of the mutants indicated that the (LSSSWT)-S-I90V and (LSSSWT)-S-Y378C AGPases have comparable allosteric and kinetic properties. However, the (LSSSWT)-S-D410G mutant exhibited altered allosteric properties of being less responsive and more sensitive to 3-phosphoglyceric acid activation and inorganic phosphate inhibition. This study not only enhances our understanding of the interaction between the SS and the LS of AGPase but also enables protein engineering to obtain enhanced assembled heat-stable variants of AGPase, which can be used for the improvement of plant yields.
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    PublicationOpen Access
    Bacterial tail anchors can target to the mitochondrial outer membrane
    (BioMed Central, 2017) Department of Molecular Biology and Genetics; Lutfullahoglu-Bal, Guleycan; Keskin, Abdurrahman; Seferoğlu, Ayşe Bengisu; Dunn, Cory David; PhD Student; Faculty Member; Department of Molecular Biology and Genetics; Graduate School of Sciences and Engineering; College of Sciences
    Background: During the generation and evolution of the eukaryotic cell, a proteobacterial endosymbiont was re-fashioned into the mitochondrion, an organelle that appears to have been present in the ancestor of all present-day eukaryotes. Mitochondria harbor proteomes derived from coding information located both inside and outside the organelle, and the rate-limiting step toward the formation of eukaryotic cells may have been development of an import apparatus allowing protein entry to mitochondria. Currently, a widely conserved translocon allows proteins to pass from the cytosol into mitochondria, but how proteins encoded outside of mitochondria were first directed to these organelles at the dawn of eukaryogenesis is not clear. Because several proteins targeted by a carboxyl-terminal tail anchor (TA) appear to have the ability to insert spontaneously into the mitochondrial outer membrane (OM), it is possible that self-inserting, tail-anchored polypeptides obtained from bacteria might have formed the first gate allowing proteins to access mitochondria from the cytosol. Results: Here, we tested whether bacterial TAs are capable of targeting to mitochondria. In a survey of proteins encoded by the proteobacterium Escherichia coli, predicted TA sequences were directed to specific subcellular locations within the yeast Saccharomyces cerevisiae. Importantly, TAs obtained from DUF883 family members ElaB and YqjD were abundantly localized to and inserted at the mitochondrial OM. Conclusions: Our results support the notion that eukaryotic cells are able to utilize membrane-targeting signals present in bacterial proteins obtained by lateral gene transfer, and our findings make plausible a model in which mitochondrial protein translocation was first driven by tail-anchored proteins.