Chemical modification of tyrosine residues in p-hydroxybenzoate hydroxylase from Pseudomonas fluorescens: assignment in sequence and catalytic involvement
- 1 July 1986
- journal article
- research article
- Published by American Chemical Society (ACS) in Biochemistry
- Vol. 25 (15), 4211-4218
- https://doi.org/10.1021/bi00363a007
Abstract
P-Hydroxybenzoate hydroxylase was modified by diethyl pyrocarbonate at pH values > 7 and by p-diazobenzoate. Modification of the enzyme by diethyl pyrocarbonate abolishes the affinity of the enzyme for the substrate p-hydroxybenzoate. Modification by p-diazobenzoate has the same effect on the enzyme. The enzyme is protected against these modifications by the effector p-fluorobenzoate. The data indicate that the modification of one tyrosine residue in the active center of the enzyme is responsible for the loss of enzyme activity. This tyrosine residue has been identified by sequence studies using radioactively labeled p-diazobenzoate and was found to be most probably Tyr-222. Diethyl pyrocarbonate reacts with a tyrosine residue in the active center other than Tyr-222; the former could not be identified. Sequence studies further showed that Cys-211 is also partially modified by p-diazobenzoate. In addition, the sequence of residues 343-345 was found to be Ser-Trp-Trp instead of the tentative assignment Ser-Tyr-Trp made earlier. The results are briefly discussed on the basis of the existing three-dimensional model of the enzyme.This publication has 21 references indexed in Scilit:
- An essential arginine residue at the substrate-binding site of p-hydroxybenzoate hydroxylase.Journal of Biological Chemistry, 1980
- PRIMARY AND TERTIARY STRUCTURE STUDIES OF PARA-HYDROXYBENZOATE HYDROXYLASE FROM PSEUDOMONAS-FLUORESCENS - ISOLATION AND ALIGNMENT OF THE CNBR PEPTIDES - INTERACTIONS OF THE PROTEIN WITH FLAVIN ADENINE-DINUCLEOTIDE1980
- A Study of p‐Hydroxybenzoate Hydroxylase from Pseudomonas fluorescensEuropean Journal of Biochemistry, 1979
- Kinetic studies on the reaction of p-hydroxybenzoate hydroxylase. Agreement of steady state and rapid reaction data.Journal of Biological Chemistry, 1979
- Crystal structure of p-hydroxybenzoate hydroxylaseJournal of Molecular Biology, 1979
- On the stable enzyme-substrate complex of p-hydroxybenzoate hydroxylase. Evidences for the proton uptake from the substrate.Journal of Biological Chemistry, 1979
- [45] Diazonium salts as specific reagents and probes of protein conformationMethods in Enzymology, 1972
- Reaction of amino acids and proteins with diazonium compounds. II. Spectra of protein derivativesArchives of Biochemistry and Biophysics, 1959
- AZOPROTEINS .1. SPECTROPHOTOMETRIC STUDIES OF AMINO ACID AZO DERIVATIVES1959
- The reactions of diazonium compounds with amino acids and proteinsBiochemical Journal, 1957