These tools will no longer be maintained as of December 31, 2024. Archived website can be found here. PubMed4Hh GitHub repository can be found here. Contact NLM Customer Service if you have questions.


Pubmed for Handhelds

PUBMED FOR HANDHELDS

Search MEDLINE/PubMed

  • Title: Order of substrate binding in bacterial phenylalanine hydroxylase and its mechanistic implication for pterin-dependent oxygenases.
    Author: Volner A, Zoidakis J, Abu-Omar MM.
    Journal: J Biol Inorg Chem; 2003 Jan; 8(1-2):121-8. PubMed ID: 12459906.
    Abstract:
    Phenylalanine hydroxylase (PAH) is a pterin-dependent non-heme metalloenzyme that catalyzes the oxidation of phenylalanine to tyrosine, which is the rate-limiting step in the catabolism of Phe. Chromobacterium violaceum phenylalanine hydroxylase (cPAH) has been prepared and its steady-state mechanism has been investigated. The enzyme requires iron for maximal activity. Initial rate measurements, done in the presence of the 6,7-dimethyl-5,6,7,8-tetrahydropterin (DMPH(4)) cofactor, yielded an average apparent k(cat) of 36+/-1 s(-1). The apparent K(M) values measured for the substrates DMPH(4), L-Phe, and O(2) are 44+/-7, 59+/-10, and 76+/-7 microM, respectively. Steady-state kinetic analyses using double-reciprocal plots revealed line patterns consistent with a sequential ter-bi mechanism in which L-Phe is the middle substrate in the order of binding. The occurrence of a line intersection on the double-reciprocal plot abscissa when either pterin or O(2) is saturated suggests that, prior to O(2) binding, DMPH(4) and L-Phe are in associative pre-equilibrium with cPAH. Together with an inhibition study using the oxidized cofactor, 7,8-dimethyl-6,7-dihydropterin, it is conclusive that the mechanism is fully ordered, with DMPH(4) binding the active site first, L-Phe second, and O(2) last. This represents the first conclusive steady-state mechanism for a PAH enzyme.
    [Abstract] [Full Text] [Related] [New Search]