• Journal of Microbiology and Biotechnology
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• v.4 no.4
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• pp.360-363
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• 1994

A colorimetric assay for $\gamma$-glutamyl transpeptidase ($\gamma$-CTP; E.C 2.3.2.2) employing 2, 4, 6-trinitrobenzene sulfonate (TNBS) to detect the amount of disappeared acceptor via transpeptidation, has been developed. Under the experimental conditions using L-$\gamma$-glutamyl ethyl ester and L-phenylalanine as $\gamma$-glutamyl donor and acceptor, respectively, it was found that the decreased absorbance of yellow color at 420 nm was strictly related to the amount of L-$\gamma$-glutamyl-L-phenylalanine (L-$\gamma$-Glu-L-Phe) formed, which was determined by DEAE-cellu-lose column chromatography. Concentrations of the enzyme and $\gamma$-glutamyl products were able to be determinedin the nanogram and nanomoles per milliliter range, respectively, with high precision and reliability. This novel assay system may therefore be a useful means for understanding of catalytic function of the $\gamma$-CTP spectrophotometrically without any usage of sophisticated instruments.

• BMB Reports
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• v.38 no.5
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• pp.609-618
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• 2005

$\gamma$-Glutamyl transpeptidase (GGT, EC 2.3.2.2.) catalyzes the transfer of the $\gamma$-glutamyl moiety from $\gamma$-glutamyl containing ompounds, notably glutathione (GSH), to acceptor amino acids and peptides. A second gene (GGTII) encoding GGT was previously isolated and characterized from the fission yeast Schizosaccharomyces pombe. In the present work, the GGTII-lacZ fusion gene was constructed and used to study the transcriptional regulation of the S. pombe GGTII gene. The synthesis of $\beta$-galactosidase from the GGTII-lacZ fusion gene was significantly enhanced by NO-generating SNP and hydrogen peroxide in the wild type yeast cells. The GGTII mRNA level was increased in the wild-type S. pombe cells treated with SNP. However, the induction by SNP was abolished in the Pap1-negative S. pombe cells, implying that the induction by SNP of GGTII is mediated by Pap1. Fermentable carbon sources, such as glucose (at low concentrations), lactose and sucrose, as a sole carbon source, enhanced the synthesis of $\beta$-galactosidase from the GGTII-lacZ fusion gene in wild type KP1 cells but not in Pap1-negative cells. Glycerol, a non-fermentable carbon source, was also able to induce the synthesis of $\beta$-galactosidase from the fusion gene, but other non-fermentable carbon sources such as acetate and ethanol were not. Transcriptional induction of the GGTII gene by fermentable carbon sources was also confirmed by increased GGTII mRNA levels in the yeast cells grown with them. Nitrogen starvation was also able to induce the synthesis of $\beta$-galactosidase from the GGTII-lacZ fusion gene in a Pap1-dependent manner. On the basis of the results, it is concluded that the S. pombe GGTII gene is regulated by oxidative and metabolic stress.

• Korean Journal of Food Science and Technology
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• v.49 no.6
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• pp.610-616
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• 2017

${\gamma}$-Glutamyltranspeptidase (GGT, EC 2.3.2.2) transfers ${\gamma}$-glutamyl moiety from glutamine to amino acids or peptides and hydrolyzes glutamine to glutamate and ammonia. In order to overproduce ${\gamma}$-glutamyltranspeptidase from Bacillus amyloliquefaciens (BAGGT), the encoding gene was cloned and expressed in Bacillus subtilis. The productivity of BAGGT in Bacillus subtilis was improved by 42-fold by using a dual-promoter system that was generated by combining promoters from B. subtilis ${\alpha}$-amylase and BAGGT genes. Through optimization of medium composition by Plackett-Burman design and central composition design, BAGGT was produced at $18.3{\times}10^7U/L$ of culture in the optimized medium. Compared to previously used Luria-Bertani medium, the optimized culture medium (15 g/L molasses, 60 g/L corn steep liquor, 6 g/L yeast extract, 4 g/L NaCl, 6 g/L $K_2HPO_4$, and 2 g/L $KH_2PO_4$), resulted in a 4.3-fold increase in production of BAGGT.