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K Seya

Publications and source records attributed to K Seya.

25 records · Page 2Linked to original sources
PubMed1994

Rapid fluorometric assay for mitochondrial proton adenosine triphosphatase activity for assessment of viability of liver graft tissue.

We developed an improved determination method of mitochondrial proton adenosine triphosphatase (ATPase) activity in the liver. The activity was measured fluorometrically with a 3,3'-dipropylthiodicarbocyanine iodide (diS-C3(5)), which is excited at 625 nm and emits fluorescence at 670 nm. This dye transmits the electric potential across the inner mitochondrial membrane. The fluorescence intensity of diS-C3(5) with mitochondria (100 microliters, 4-16 mg/ml protein) in a 2 ml potassium buffer (pH 7.4) was regarded as a standard electric potential. After confirming the activity of the mitochondrial electron transport chain by succinic acid (9 mumol), we inhibited the chain by antimycin A (1.25 micrograms). Fluorescence intensity decreased by adenosine 5'-triphosphate (ATP) (2 mumol) and oligomycin (25 micrograms) inhibited this depression. The value of mitochondrial proton ATPase activity was calculated as a percentage of the fluorescence intensity change by ATP per the standard electric potential. The activity of mitochondrial proton ATPase in the normal fresh rat livers was 50.3 +/- 2.2%. Good correlation (r2 = 0.807) between two methods for mitochondrial proton ATPase activity, our newly developed method and a conventional colorimetric method, was obtained in the rat livers with various conditions. This method has advantages that the proton ATPase activity can be measured in intact mitochondria, and all procedures can be completed within 40 min. It is suitable for the determination of mitochondrial viability of liver graft in the hepatic resections and transplantations.

Adenosine Triphosphate↗
PubMed1988

Physicochemical properties of charge isomers of recombinant human superoxide dismutase.

Recombinant human Cu2Zn2SOD expressed in Escherichia coli consisted of mainly three isomers with isoelectric points of 5.14 (A), 5.06 (B), and 4.99 (C). Each isomer was isolated by DEAE-Toyopearl chromatography and the physiochemical properties were investigated. No significant differences in chemical and spectrophotometric properties, such as specific activity, metal contents, amino acid composition, and UV and ESR spectra, were found. The result of labeling of free cysteine residues with ABD-F showed the disulfide bond to be formed between 57Cys and 146Cys in every isomer. A few differences were found in the CD spectrum around 260 nm and in the elution patterns on reverse-phase HPLC. The isoelectric points of the three isomers became the same after treatment by reduction and carboxymethylation and even after reduction only, pI of isomers tended to be at the value of component (A). These results suggest that the three isomers are identical in primary structure but slightly different in secondary or tertiary structure. These differences are probably derived from structural alterations around 111Cys.

Amino Acids↗
PubMed1987

Synthesis of p-hydroxyubenimex.

p-Hydroxyubenimex, (2S,3R)-3-amino-2-hydroxy-4-p-hydroxyphenylbutyryl-L-leucine, was synthesized starting from D-tyrosine. The structure and stereochemistry of the synthesized product were confirmed by comparison with p-hydroxyubenimex that was chemically transformed from ubenimex, an aminopeptidase inhibitor of microbial origin. Compared to ubenimex, p-hydroxyubenimex is more active against aminopeptidase B but less active against leucine aminopeptidase. By using the synthetic p-hydroxyubenimex as a reference sample, one of the metabolites of ubenimex was identified as p-hydroxyubenimex. The (2R,3R)-stereoisomer of p-hydroxyubenimex was also prepared. However, its activity against aminopeptidases was much weaker.

Aminopeptidases↗