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    Comparative modelling of a novel enzyme: Mus musculus leucine decarboxylase
    (Tubitak Scientific & Technological Research Council Turkey, 2020) Sahutoglu, Arif Sercan
    Leucine decarboxylase (LDC) is a recently proposed enzyme with no official enzyme commission number yet. It is encoded by the Mus musculus gene Gm853 which is expressed at kidneys, generating isopentylamine, an alkylmonoamine that has not been described to be formed by any metazoan enzyme yet. Although the relevance of LDC in mammalian physiology has not been fully determined, isopentylamine is a potential modulator which may have effects on insulin secretion and healthy gut microbiota formation. The LDC is a stable enzyme that specifically decarboxylates L-leucine but does not decarboxylate ornithine or lysine as its paralogues ornithine decarboxylase (ODC; EC: 4.1.1.17) and lysine decarboxylase (KDC; EC: 4.1.1.18) do. It does not act as an antizyme inhibitor and does not decarboxylate branched amino acids such as valine and isoleucine as it is another paralogue valine decarboxylase (VDC; EC: 4.1.1.14). The crystal structure of the enzyme has not been determined yet but there are homologous structures with complete coverage in Protein Data Bank (PDB) which makes LDC a good candidate for comparative modelling. In this study, homology models of LDC were generated and used in cofactor and substrate docking to understand the structure/function relationship underlying the unique selectivity of LDC enzyme.
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    Fine-tuned preparation of cross-linked laccase nanoaggregates
    (Taylor & Francis Ltd, 2019) Sahutoglu, Arif Sercan; Akgul, Cahit
    This study focuses on well-known but commonly overlooked or unreported factors in the preparation of cross-linked enzyme nano-aggregates (nano-CLEAs). The parameters including the ionic strength of the protein solution, protein, precipitant and cross-linker concentrations, pH and addition order of the reagents were fine-tuned for nanoaggregate preparation without the need of non-protein support material, special equipment or sophisticated procedures. For this purpose, precipitation as nano-aggregates and then cross-linking while maintaining submicron size distribution were studied independently. Moreover, nano-aggregate formation from reverse micellar solutions was also investigated to improve the scope of the method to membrane-bound enzymes. Five different precipitation agents together with three different cross-linkers were investigated for immobilization of the Trametes versicolor laccase as cross-linked nano-aggregates. Although complete activity recovery was possible for micro-aggregates, the best activity results for nano-aggregates were 40.5%+/- 5.0. The K-m values of the immobilized enzymes were slightly lower than the K-m values of the free counterparts which indicate little or no mass transfer limitation due to the nano-immobilization process. However, V-max values were also lower. The activity loss and V-max reduction upon immobilization were found to mainly result from the modification of the amine groups instead of excess crosslinking. The thermal stabilities of the crosslinked laccase nano-aggregates were significantly higher (similar to 10-30 fold at 60 degrees C) compared to free laccase and the nano-CLEAs retained up to 30% of their initial activities upon 7 consequent usages. The sizes of the obtained immobilized enzyme products were found to be greatly variable depending on the cross-linker type. The smaller particles (similar to 200 nm radius) were obtained when EDAC was used as the cross-linker. The larger products (similar to 600 nm radius) were prepared when the cross-linker was dextran poly-aldehyde. The addition order of the reagents was found to be effective on particle size and thermal stability values.
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    Isolation of ethanol-tolerant Saccharomyces cerevisiae strains by using a novel mutagenesis method
    (Elsevier Sci Ltd, 2011) Pazarlioglu, Nurdan Kasikara; Ensari, Yunus; Sahutoglu, Arif Sercan
    [Anstract Not Available]
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    Lactoferrin for COVID-19 prevention, treatment, and recovery
    (Frontiers Media Sa, 2022) Bolat, Ecem; Eker, Furkan; Kaplan, Merve; Duman, Hatice; Arslan, Aysenur; Saritas, Suemeyye; Sahutoglu, Arif Sercan
    Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2), a unique beta-coronavirus, has caused the most serious outbreak of the last century at the global level. SARS-CoV-2 infections were firstly reported in the city of Wuhan in China in 2019 and this new disease was named COVID-19 by World Health Organization (WHO). As this novel disease can easily be transmitted from one individual to another via respiratory droplets, many nations around the world have taken several precautions regarding the reduction in social activities and quarantine for the limitation of the COVID-19 transmission. SARS-CoV-2 is known to cause complications that may include pneumonia, acute respiratory distress syndrome (ARDS), multi-organ failure, septic shock, and death. To prevent and treat COVID-19, some significant studies have been conducted since the outbreak. One of the most noticeable therapeutic approaches is related to a multifunctional protein, lactoferrin. Lactoferrin (Lf) is an 80 kDa cationic glycoprotein that has a great range of benefits from improving the immunity to antiviral effects due to its unique characteristics such as the iron-binding ability. This review summarizes the characteristics of SARS-CoV-2 and the potential applications of Lf for the prevention, treatment, and recovery of COVID-19.
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    One-phase synthesis of single enzyme nanoparticles (SENs) of Trametes versicolor laccase by in situ acrylamide polymerisation
    (Taylor & Francis Ltd, 2020) Sahutoglu, Arif Sercan; Akgul, Cahit
    Single enzyme nanoparticles (SENs) are a state of art technology that allows immobilisation of individual enzyme molecules in a nanometre scale cross-linked polymer matrix that is covalently bonded to the enzyme. Although the method has great potential in the enzyme immobilisation, it has attracted relatively low attention from researchers over the last decade. Therefore, it can be very fruitful to apply this method to several enzymes in order to increase the understanding about the mechanisms and parameters important for the technique. In this study, Trametes versicolor laccase enzyme (TvL) was immobilised as SENs without phase transfer. The enzyme was immobilised via acryloyl chloride modification that is followed by an in situ polyacrylamide polymerisation in the water phase. The resulting single TvL nanoparticles were found to be less than 50 nm in diameter and round in shape with an activity recovery of 66.33 +/- 2.57%. Both V-max and K-m values of the TvL SENs were lower than the free counterpart. However, approximately the same K-cat/K-m values suggested similar catalytic efficiencies for the free and the immobilised enzymes. The significantly lower (similar to 2 fold) K-m value of the immobilised enzyme suggested an affinity increase towards 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonate) (ABTS) upon immobilisation. Whilst the immobilised and the free enzymes showed similar thermal stabilities at 60 degrees C for 240 min, the immobilised form showed higher stability in solution at ambient temperature after 15 days which may suggest an increase in microbial stability.
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    Role of milk glycome in prevention, treatment, and recovery of COVID-19
    (Frontiers Media Sa, 2022) Kaplan, Merve; Sahutoglu, Arif Sercan; Saritas, Sumeyye; Duman, Hatice; Arslan, Aysenur; Pekdemir, Burcu; Karav, Sercan
    Milk contains all essential macro and micro-nutrients for the development of the newborn. Its high therapeutic and antimicrobial content provides an important function for the prevention, treatment, and recovery of certain diseases throughout life. The bioactive components found in milk are mostly decorated with glycans, which provide proper formation and modulate the biological functions of glycosylated compounds. The glycome of milk consists of free glycans, glycolipids, and N- and O- glycosylated proteins. Recent studies have shown that both free glycans and glycan-containing molecules have antiviral characteristics based on different mechanisms such as signaling, microbiome modulation, natural decoy strategy, and immunomodulatory action. In this review, we discuss the recent clinical studies and potential mechanisms of free and conjugated glycans' role in the prevention, treatment, and recovery of COVID-19.
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    Structural insights of two novel N-acetyl-glucosaminidase enzymes through in silico methods
    (Tubitak Scientific & Technological Research Council Turkey, 2020) Sahutoglu, Arif Sercan; Duman, Hatice; Frese, Steven Alex; Karav, Sercan
    EndoBI-1 and EndoBI-2 are two endo-beta-N-acetylglucosaminidase isoenzymes that cleave N-N'-diacetylchitobiosyl moieties found in various types of native N-glycans. These N-glycans are indigestible by human infants and adults due to the lack of responsible glycosyl hydrolases and they act as selective prebiotics for a probiotic microorganism, Bifidobacterium longum subsp. infantis, in the large intestine. The selectivity and the thermostability of EndoBI-1 and EndoBI-2 suggest that these enzymes may be useful for many scientific and industrial applications. In this study, the growing numbers of homologous sequences in different databases were exploited in a comparative approach to investigate structural properties of EndoBI-1 and EndoBI-2 enzymes. Moreover, the complete and partial homology models of these two enzymes were generated and evaluated. Selected models were used for docking studies of the plus subsite ligand of these enzymes for further understanding on the substrate selectivity of EndoBI enzymes.
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    Synthesis, characterization, carbonic anhydrase inhibitor activity, and docking studies of phenylthiazol-2 (3h)-ylidene-isoquinoline-5-amine Derivatives
    (Elsevier, 2023) Berber, Nurcan; Sahutoglu, Arif Sercan; Gokce, Basak; Cikrikci, Kuebra; Gencer, Nahit
    Phenylthiazol-2(3h)-ylidene-isoquinoline-5-amine derivatives(5a-r) were easily prepared from 5-aminoisoquinoline, thioisocyanate, and phenacyl bromide derivatives in the presence of triethylamine in tetrahydrofuran (THF), and dimethyl formamide (DMF) in two steps. In the first step, thiourea was synthesized in THF reflux, 24 h and then in the second step, the formation of thiazole ring was ensured in EtOH-DMF (5:5 v/v), reflux, 24 h. The newly synthesized compounds were characterized using 1H NMR, 13C NMR, IR, and elemental analysis. The inhibitory effects of 18 newly synthesized compounds (5a-r) on hydratase and esterase activities of carbonic anhydrase isoenzymes (hCA I, II, IX, and X) were studied in vitro.