Staff Publications

Staff Publications

  • external user (warningwarning)
  • Log in as
  • language uk
  • About

    'Staff publications' is the digital repository of Wageningen University & Research

    'Staff publications' contains references to publications authored by Wageningen University staff from 1976 onward.

    Publications authored by the staff of the Research Institutes are available from 1995 onwards.

    Full text documents are added when available. The database is updated daily and currently holds about 240,000 items, of which 72,000 in open access.

    We have a manual that explains all the features 

Record number 445968
Title The Reaction Kinetics of 3-Hydroxybenzoate 6-Hydroxylase from Rhodococcus jostii RHA1 Provide an Understanding of the para-Hydroxylation Enzyme Catalytic Cycle
Author(s) Sucharitakul, J.; Tongsook, C.; Pakotiprapha, D.; Berkel, W.J.H. van; Chaiyen, P.
Source Journal of Biological Chemistry 288 (2013)49. - ISSN 0021-9258 - p. 35210 - 35221.
Department(s) Biochemistry
Publication type Refereed Article in a scientific journal
Publication year 2013
Keyword(s) para-hydroxybenzoate hydroxylase - p-hydroxyphenylacetate 3-hydroxylase - 2-methyl-3-hydroxypyridine-5-carboxylic acid oxygenase - steady-state - ornithine hydroxylase - vibrio-campbellii - crystal-structure - gentisic acid - in-vitro - mechanism
Abstract 3-Hydroxybenzoate 6-hydroxylase (3HB6H) from Rhodococcus jostii RHA1 is an NADH-specific flavoprotein monooxygenase that catalyzes the para-hydroxylation of 3-hydroxybenzoate (3HB) to form 2,5-dihydroxybenzoate (2,5-DHB). Based on results from stopped-flow spectrophotometry, the reduced enzyme-3HB complex reacts with oxygen to form a C4a-peroxy flavin with a rate constant of 1.13 ± 0.01 × 10(6) m(-1) s(-1) (pH 8.0, 4 °C). This intermediate is subsequently protonated to form a C4a-hydroperoxyflavin with a rate constant of 96 ± 3 s(-1). This step shows a solvent kinetic isotope effect of 1.7. Based on rapid-quench measurements, the hydroxylation occurs with a rate constant of 36 ± 2 s(-1). 3HB6H does not exhibit substrate inhibition on the flavin oxidation step, a common characteristic found in most ortho-hydroxylation enzymes. The apparent kcat at saturating concentrations of 3HB, NADH, and oxygen is 6.49 ± 0.02 s(-1). Pre-steady state and steady-state kinetic data were used to construct the catalytic cycle of the reaction. The data indicate that the steps of product release (11.7 s(-1)) and hydroxylation (36 ± 2 s(-1)) partially control the overall turnover
There are no comments yet. You can post the first one!
Post a comment
Please log in to use this service. Login as Wageningen University & Research user or guest user in upper right hand corner of this page.