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349 a.a.
__K
_NA
Bound ligand (Het Group name = PHE) corresponds exactly |
+ |
H(2)O |
+ |
 NAD(+) |
= |
 phenylpyruvate |
+ |
NH(3) |
+ |
Bound ligand (Het Group name = NAD) corresponds exactly |
|---|
Key reference
DOI no: 10.1021/bi000494c Biochemistry 39:9174-9187 (2000) PubMed id: 10924111
Rhodococcus L-phenylalanine dehydrogenase: kinetics, mechanism, and structural basis for catalytic specificity. N.M.Brunhuber, J.B.Thoden, J.S.Blanchard, J.L.Vanhooke. ABSTRACT
Phenylalanine dehydrogenase catalyzes the reversible, pyridine nucleotide-dependent oxidative deamination of L-phenylalanine to form phenylpyruvate and ammonia. We have characterized the steady-state kinetic behavior of the enzyme from Rhodococcus sp. M4 and determined the X-ray crystal structures of the recombinant enzyme in the complexes, E.NADH.L-phenylalanine and E.NAD(+). L-3-phenyllactate, to 1.25 and 1.4 A resolution, respectively. Initial velocity, product inhibition, and dead-end inhibition studies indicate the kinetic mechanism is ordered, with NAD(+) binding prior to phenylalanine and the products' being released in the order of ammonia, phenylpyruvate, and NADH. The enzyme shows no activity with NADPH or other 2'-phosphorylated pyridine nucleotides but has broad activity with NADH analogues. Our initial structural analyses of the E.NAD(+).phenylpyruvate and E.NAD(+). 3-phenylpropionate complexes established that Lys78 and Asp118 function as the catalytic residues in the active site [Vanhooke et al. (1999) Biochemistry 38, 2326-2339]. We have studied the ionization behavior of these residues in steady-state turnover and use these findings in conjunction with the structural data described both here and in our first report to modify our previously proposed mechanism for the enzymatic reaction. The structural characterizations also illuminate the mechanism of the redox specificity that precludes alpha-amino acid dehydrogenases from functioning as alpha-hydroxy acid dehydrogenases.
Literature references that cite this PDB file's key reference
PubMed id Reference
12192068 C.A.Bottoms, P.E.Smith, and J.J.Tanner (2002).
A structurally conserved water molecule in Rossmann dinucleotide-binding domains.Protein Sci, 11, 2125-2137.
12220195 J.F.Tally, S.J.Maniscalco, S.K.Saha, and H.F.Fisher (2002).
Detection of multiple active site domain motions in transient-state component time courses of the Clostridium symbiosum L-glutamate dehydrogenase-catalyzed oxidative deamination reaction.Biochemistry, 41, 11284-11293.
12037317 T.A.Muranova, S.N.Ruzheinikov, S.E.Sedelnikova, P.J.Baker, A.Pasquo, A.Galkin, N.Esaki, T.Ohshima, K.Soda, and D.W.Rice (2002).
Crystallization and preliminary X-ray analysis of substrate complexes of leucine dehydrogenase from Thermoactinomyces intermedius.Acta Crystallogr D Biol Crystallogr, 58, 1059-1062. The most recent references are shown first. Citation data come partly from CiteXplore and partly from an automated harvesting procedure. Note that this is likely to be only a partial list as not all journals are covered by either method. However, we are continually building up the citation data so more and more references will be included with time.
