Molecular Biology and Genetics / Moleküler Biyoloji ve Genetik
Permanent URI for this collectionhttps://hdl.handle.net/11147/9
Browse
7 results
Search Results
Article Citation - WoS: 16Citation - Scopus: 15Gene Cloning, Heterologous Expression, and Partial Characterization of a Novel Cold-Adapted Subfamily I.3 Lipase From Pseudomonas Fluorescence Ke38(Nature Publishing Group, 2020) Karakaş, Fulya; Arslanoğlu, AlperA novel cold-active true lipase from Pseudomonas sp. KE38 was cloned, sequencing and expressed in E. coli by degenerate PCR and genome walking technique. The open reading frame of the cloned gene encoded a polypeptide chain of 617 amino acids with a confirmed molecular weight of 64 kD. Phylogenetic analysis of the deduced amino acid sequence of the lipase indicated that it had high similarity with lipases of subfamily ?.3 of bacterial lipases. Recombinant lipase was purified in denatured form as inclusion bodies, which were then renatured by urea followed by dialysis. Lipase activity was determined titrimetrically using olive oil as substrate. The enzyme showed optimal activity at 25 °C, pH 8.5 and was highly stable in the presence of various metal ions and organic solvents. Low optimal temperature and high activity in the presence of methanol and ethanol make this lipase a potential candidate for transesterification reactions and biodiesel production. © 2020, The Author(s).Conference Object Keratitis-Ichthyosis Syndrome Associated Mutations Impair the Localization and Functions of Connexin 26(Nature Publishing Group, 2015) Aypek, Hande; Meşe, GülistanConnexins (Cx) form gap junctions and non-junctional hemichannels that play roles in several cellular mechanisms, including proliferation and differentiation. The importance of connexins for human physiology was shown by the association of mutations in several isoforms with various human hereditary disorders. Mutations in Cx26 cause both non-syndromic and syndromic deafness associated with skin disorders including keratitis-ichthyosis-deafness (KID) syndrome. In vitro characterization of Cx26 mutations suggested that mutations causing non-syndromic deafness and syndromic deafness show different properties, where the former ones result in loss-of-function and the latter ones cause gain-of-function mutations.Article Citation - WoS: 79Citation - Scopus: 94Biofabrication of in Situ Self Assembled 3d Cell Cultures in a Weightlessness Environment Generated Using Magnetic Levitation(Nature Publishing Group, 2018) Anıl İnevi, Müge; Yaman, Sena; Arslan Yıldız, Ahu; Meşe, Gülistan; Yalçın Özuysal, Özden; Tekin, Hüseyin Cumhur; Özçivici, EnginMagnetic levitation though negative magnetophoresis is a novel technology to simulate weightlessness and has recently found applications in material and biological sciences. Yet little is known about the ability of the magnetic levitation system to facilitate biofabrication of in situ three dimensional (3D) cellular structures. Here, we optimized a magnetic levitation though negative magnetophoresis protocol appropriate for long term levitated cell culture and developed an in situ 3D cellular assembly model with controlled cluster size and cellular pattern under simulated weightlessness. The developed strategy outlines a potential basis for the study of weightlessness on 3D living structures and with the opportunity for real-time imaging that is not possible with current ground-based simulated weightlessness techniques. The low-cost technique presented here may offer a wide range of biomedical applications in several research fields, including mechanobiology, drug discovery and developmental biology.Article Citation - WoS: 37Citation - Scopus: 45On the Performance of Pre-Microrna Detection Algorithms(Nature Publishing Group, 2017) Saçar Demirci, Müşerref Duygu; Baumbach, Jan; Allmer, JensMicroRNAs are crucial for post-transcriptional gene regulation, and their dysregulation has been associated with diseases like cancer and, therefore, their analysis has become popular. The experimental discovery of miRNAs is cumbersome and, thus, many computational tools have been proposed. Here we assess 13 ab initio pre-miRNA detection approaches using all relevant, published, and novel data sets while judging algorithm performance based on ten intrinsic performance measures. We present an extensible framework, izMiR, which allows for the unbiased comparison of existing algorithms, adding new ones, and combining multiple approaches into ensemble methods. In an exhaustive attempt, we condense the results of millions of computations and show that no method is clearly superior; however, we provide a guideline for biomedical researchers to select a tool. Finally, we demonstrate that combining all of the methods into one ensemble approach, for the first time, allows reliable purely computational pre-miRNA detection in large eukaryotic genomes.Article Citation - WoS: 45Citation - Scopus: 48Connexin26 Mutations Causing Palmoplantar Keratoderma and Deafness Interact With Connexin43, Modifying Gap Junction and Hemichannel Properties(Nature Publishing Group, 2016) Shuja, Zunaira; Li, Leping; Gupta, Shashank; Meşe, Gülistan; White, Thomas W.Mutations in GJB2 (connexin [Cx]26) cause either deafness or deafness associated with skin diseases. That different disorders can be caused by distinct mutations within the same gene suggests that unique channel activities are influenced by each class of mutation. We have examined the functional characteristics of two human mutations, Cx26-H73R and Cx26-S183F, causing palmoplantar keratoderma (PPK) and deafness. Both failed to form gap junction channels or hemichannels when expressed alone. Coexpression of the mutants with wild-type Cx43 showed a transdominant inhibition of Cx43 gap junction channels, without reductions in Cx43 protein synthesis. In addition, the presence of mutant Cx26 shifted Cx43 channel gating and kinetics toward a more Cx26-like behavior. Coimmunoprecipitation showed Cx43 being pulled down more efficiently with mutant Cx26 than wild-type, confirming the enhanced formation of heteromeric connexons. Finally, the formation of heteromeric connexons resulted in significantly increased Cx43 hemichannel activity in the presence of Cx26 mutants. These findings suggest a common mechanism whereby Cx26 mutations causing PPK and deafness transdominantly influence multiple functions of wild-type Cx43. They also implicate a role for aberrant hemichannel activity in the pathogenesis of PPK and further highlight an emerging role for Cx43 in genetic skin diseases. © 2015 The Authors. Published by Elsevier, Inc. on behalf of the Society for Investigative Dermatology.Article Citation - WoS: 49Citation - Scopus: 495-Fluorouracil Signaling Through a Calcium-Calmodulin Pathway Is Required for P53 Activation and Apoptosis in Colon Carcinoma Cells(Nature Publishing Group, 2013) Can, G.; Akpınar, B.; Baran, Yusuf; Zhivotovsky, B.; Olsson, M.5-Fluorouracil (5-FU) is an anti-metabolite that is in clinical use for treatment of several cancers. In cells, it is converted into three distinct fluoro-based nucleotide analogs, which interfere with DNA synthesis and repair, leading to genome impairment and, eventually, apoptotic cell death. Current knowledge states that in certain cell types, 5-FU-induced stress is signaling through a p53-dependent induction of tumor necrosis factor-receptor oligomerization required for death-inducing signaling complex formation and caspase-8 activation. Here we establish a role of calcium (Ca 2+) as a messenger for p53 activation in response to 5-FU. Using a combination of pharmacological and genetic approaches, we show that treatment of colon carcinoma cells stimulates entry of extracellular Ca 2+ through long lasting-type plasma membrane channels, which further directs posttranslational phosphorylation of at least three p53 serine residues (S15, S33 and S37) by means of calmodulin (CaM) activity. Obstructing this pathway by the Ca 2+ -chelator BAPTA (1,2-bis(o-aminophenoxy)ethane- N,N,N',N'-tetraacetic acid) or by inhibitors of CaM efficiently reduces 5-FU-induced caspase activities and subsequent cell death. Moreover, ectopic expression of p53 S15A in HCT116 p53 -/- cells confirmed the importance of a Ca 2+ -CaM-p53 axis in 5-FU-induced extrinsic apoptosis. The fact that a widely used therapeutic drug, such as 5-FU, is operating via this pathway could provide new therapeutic intervention points, or specify new combinatorial treatment regimes. © 2013 Macmillan Publishers Limited.Article Citation - WoS: 133Citation - Scopus: 138Antagonistic Roles of Notch and P63 in Controlling Mammary Epithelial Cell Fates(Nature Publishing Group, 2010) Yalçın Özuysal, Özden; Fiche, M.; Guitierrez, M.; Wagner, K. U.; Raffoul, W.; Brisken, C.The breast epithelium has two major compartments, luminal and basal cells, that are established and maintained by poorly understood mechanisms. The p53 homolog, p63, is required for the formation of mammary buds, but its function in the breast after birth is unknown. We show that in primary human breast epithelial cells, maintenance of basal cell characteristics depends on continued expression of the p63 isoform, ΔNp63, which is expressed in the basal compartment. Forced expression of ΔNp63 in purified luminal cells confers a basal phenotype. Notch signaling downmodulates ΔNp63 expression and mimics ΔNp63 depletion, whereas forced expression of ΔNp63 partially counteracts the effects of Notch. Consistent with Notch activation specifying luminal cell fate in the mammary gland, Notch signaling activity is specifically detected in mice at sites of pubertal ductal morphogenesis where luminal cell fate is determined. Basal cells in which Notch signaling is active show decreased p63 expression. Both constitutive expression of ΔNp63 and ablation of Notch signaling are incompatible with luminal cell fate. Thus, the balance between basal and luminal cell compartments of the breast is regulated by antagonistic functions of ΔNp63 and Notch. © 2010 Macmillan Publishers Limited All rights reserved.