PubMed:10441144
Annnotations
{"target":"https://pubannotation.org/docs/sourcedb/PubMed/sourceid/10441144","sourcedb":"PubMed","sourceid":"10441144","source_url":"http://www.ncbi.nlm.nih.gov/pubmed/10441144","text":"Spectral and kinetic studies of the oxidation of monosubstituted phenols and anilines by recombinant Synechocystis catalase-peroxidase compound I.\nA high-level expression in Escherichia coli of a fully active recombinant form of a catalase-peroxidase (KatG) from the cyanobacterium Synechocystis PCC 6803 is reported. Since both physical and kinetic characterization revealed its identity with the wild-type protein, the large quantities of recombinant KatG allowed the first examination of second-order rate constants for the oxidation of a series of aromatic donor molecules (monosubstituted phenols and anilines) by a bifunctional catalase-peroxidase compound I using the sequential-mixing stopped-flow technique. Because of the overwhelming catalase activity, peroxoacetic acid has been used for compound I formation. A \u003e/=50-fold excess of peroxoacetic acid is required to obtain a spectrum of relatively pure and stable compound I which is characterized by about 40% hypochromicity, a Soret maximum at 406 nm, and isosbestic points between the native enzyme and compound I at 357 and 430 nm. The apparent second-order rate constant for formation of compound I from ferric enzyme and peroxoacetic acid is (8.74 +/- 0.26) x 10(3) M(-)(1) s(-)(1) at pH 7. 0. Reduction of compound I by aromatic donor molecules is dependent upon the substituent effect on the benzene ring. The apparent second-order rate constants varied from (3.6 +/- 0.1) x 10(6) M(-)(1) s(-)(1) for p-hydroxyaniline to (5.0 +/- 0.1) x 10(2) M(-)(1) s(-)(1) for p-hydroxybenzenesulfonic acid. They are shown to correlate with the substituent constants in the Hammett equation, which suggests that in bifunctional catalase-peroxidases the aromatic donor molecule donates an electron to compound I and loses a proton simultaneously. The value of rho, the susceptibility factor in the Hammett equation, is -3.4 +/- 0.4 for the phenols and -5.1 +/- 0.8 for the anilines. The pH dependence of compound I reduction by aniline exhibits a relatively sharp maximum at pH 5. The redox intermediate formed upon reduction of compound I has spectral features which indicate that the single oxidizing equivalent in KatG compound II is contained on an amino acid which is not electronically coupled to the heme.","tracks":[{"project":"CyanoBase","denotations":[{"id":"T1","span":{"begin":115,"end":134},"obj":"protein"},{"id":"T2","span":{"begin":231,"end":250},"obj":"protein"},{"id":"T3","span":{"begin":252,"end":256},"obj":"protein"},{"id":"T4","span":{"begin":453,"end":457},"obj":"protein"},{"id":"T5","span":{"begin":634,"end":653},"obj":"protein"},{"id":"T6","span":{"begin":1684,"end":1704},"obj":"protein"},{"id":"T7","span":{"begin":2172,"end":2176},"obj":"protein"}],"attributes":[{"subj":"T1","pred":"source","obj":"CyanoBase"},{"subj":"T2","pred":"source","obj":"CyanoBase"},{"subj":"T3","pred":"source","obj":"CyanoBase"},{"subj":"T4","pred":"source","obj":"CyanoBase"},{"subj":"T5","pred":"source","obj":"CyanoBase"},{"subj":"T6","pred":"source","obj":"CyanoBase"},{"subj":"T7","pred":"source","obj":"CyanoBase"}]}],"config":{"attribute types":[{"pred":"source","value type":"selection","values":[{"id":"CyanoBase","color":"#d993ec","default":true}]}]}}