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Biomedical subjects

Y Saeki

Publications and source records attributed to Y Saeki.

At least 271 records · Page 15Linked to original sources
PubMed1976

Nonidentical subunits of protocatechuate 3,4-dioxygenase.

Protocatechuate 3,4-dioxygenase (EC 1.13.11.3) has been reported to have a molecular weight of 700,000 and to consist of eight identical subunits, each containing one atom of ferric iron and a substrate binding site. This subunit has now been found to dissociate further into four smaller subunits of two nonidentical types (alpha2beta2), upon sodium dodecyl sulfate gel electrophoresis. The molecular weights of the alpha and beta subunits were estimated to be 22,500 and 25,000, respectively. Isoelectric focusing of the enzyme in 6 M urea revealed that the isoelectric points of the alpha and beta subunits were 5.2 and 9.5, respectively. Separation of the two subunits was achieved by chromatography on sulfopropyl (SP)-Sephadex in 6 M urea after treatment of the enzyme with 8 M urea at 37 degrees C for 6 h. The NH2-terminal sequence of the alpha subunit was determined to be Pro-Ile-Glu-Leu-Leu-Pro-Glu-Thr-Pro-Ser-Glx-Thr-Ala-Gly and that of the beta subunit, Pro-Ala-Gln-Asp-Asn-Ala-Arg-Phe-Val-Ile-Arg-Asx-Arg-Asx. Phenylalanine was found as the COOH-terminal residue of the alpha subunit. However, the COOH terminus of the beta subunit was not detected by any of three methods employed.

Amino Acid Sequence↗
PubMed1975

Extradiol cleavage of 3-substituted catechols by an intradiol dioxygenase, pyrocatechase, from a Pseudomonad.

Pyrocatechase (catechol 1,2-oxidoreductase (decyclizing), EC 1.13.11.1), a ferric ion-containing dioxygenase from Pseudomonas arvilla C-1, catalyzes the intradiol cleavage of catechol with insertion of 2 atoms of molecular oxygen to form cis,cis-muconic acid. The enzyme also catalyzed the oxidation of various catechol derivatives, including 4-methylcatechol, 4-chlorocatechol, 4-formylcatechol (protocatechualdehyde), 4,5-dichlorocatechol, 3,5-dichlorocatechol, 3-methylcatechol, 3-methoxycatechol, and 3-hydroxycatechol (pyrogallol). All of these substrates gave products having an absorption maximum at around 260 nm, which is characteristic of cis,cis-muconic acid derivatives. However, when 3-methylcatechol was used as substrate, the product formed showed two absorption maxima at 390 and 260 nm. These two absorption maxima were found to be attributable to two different products, 2-hydroxy-6-oxo-2,4-heptadienoic acid and 5-carboxy-2-methyl-2,4-pentadienoic acid (2-methylmuconic acid). The former was produced by the extradiol cleavage between the carbon atom carrying the hydroxyl group and the carbon atom carrying the hydroxyl group and the carbon atom carrying the methyl group; the latter by an intradiol cleavage between two hydroxyl groups. Since these products were unstable, they were converted to and identified as 6-methylpyridine-2-carboxylic acid and 2-methylmuconic acid dimethylester, respectively. Similarly, 3-methoxycatechol gave two products, namely, 2-hydroxy-5-methoxycarbonyl-2,4-pentadienoic acid and 5-carboxy-2-methoxy-2,4-pentadienoic acid (2-methoxymuconic acid). With 3-methylcatechol as substrate, the ratio of intradiol and extradiol cleavage activities of Pseudomonas pyrocatechase during purification was almost constant and was about 17. The final preparation of the enzyme was homogeneous when examined by disc gel electrophoresis and catalyzed both reactions simultaneously with the same ratio as during purification. All attempts to resolve the enzyme into two components with separate activities, including inactivation of the enzyme with urea or heat, treatment with sulfhydryl-blocking reagents or chelating agents, and inhibition of the enzyme with various inhibitors, proved unsuccessful. These results strongly suggest that Pseudomonas pyrocatechase is a single enzyme, which catalyzes simultaneously both intradiol and extradiol cleavages of some 3-substituted catechols.

1,2-Dihydroxybenzene-3,5-Disulfonic Acid Disodium ↗