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A core catalytic domain of the TyrA protein family: arogenate dehydrogenase from Synechocystis.

ABSTRACT: The TyrA protein family includes prephenate dehydrogenases, cyclohexadienyl dehydrogenases and TyrA(a)s (arogenate dehydrogenases). tyrA(a) from Synechocystis sp. PCC 6803, encoding a 30 kDa TyrA(a) protein, was cloned into an overexpression vector in Escherichia coli. TyrA(a) was then purified to apparent homogeneity and characterized. This protein is a model structure for a catalytic core domain in the TyrA superfamily, uncomplicated by allosteric or fused domains. Competitive inhibitors acting at the catalytic core of TyrA proteins are analogues of any accepted cyclohexadienyl substrate. The homodimeric enzyme was specific for L-arogenate (K(m)=331 microM) and NADP+ (K(m)=38 microM), being unable to substitute prephenate or NAD+ respectively. L-Tyrosine was a potent inhibitor of the enzyme (K(i)=70 microM). NADPH had no detectable ability to inhibit the reaction. Although the mechanism is probably steady-state random order, properties of 2',5'-ADP as an inhibitor suggest a high preference for L-arogenate binding first. Comparative enzymology established that both of the arogenate-pathway enzymes, prephenate aminotransferase and TyrA(a), were present in many diverse cyanobacteria and in a variety of eukaryotic red and green algae.

SUBMITTER: Bonner CA

PROVIDER: S-EPMC1133941 | biostudies-literature | 2004 Aug

REPOSITORIES: biostudies-literature

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A core catalytic domain of the TyrA protein family: arogenate dehydrogenase from Synechocystis.

Bonner Carol A CA Jensen Roy A RA Gander John E JE Keyhani Nemat O NO

The Biochemical journal 20040801 Pt 1

The TyrA protein family includes prephenate dehydrogenases, cyclohexadienyl dehydrogenases and TyrA(a)s (arogenate dehydrogenases). tyrA(a) from Synechocystis sp. PCC 6803, encoding a 30 kDa TyrA(a) protein, was cloned into an overexpression vector in Escherichia coli. TyrA(a) was then purified to apparent homogeneity and characterized. This protein is a model structure for a catalytic core domain in the TyrA superfamily, uncomplicated by allosteric or fused domains. Competitive inhibitors actin ...[more]

PMID: 15171683

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Project description:BackgroundThe TyrA protein family includes members that catalyze two dehydrogenase reactions in distinct pathways leading to L-tyrosine and a third reaction that is not part of tyrosine biosynthesis. Family members share a catalytic core region of about 30 kDa, where inhibitors operate competitively by acting as substrate mimics. This protein family typifies many that are challenging for bioinformatic analysis because of relatively modest sequence conservation and small size.ResultsPhylogenetic relationships of TyrA domains were evaluated in the context of combinatorial patterns of specificity for the two substrates, as well as the presence or absence of a variety of fusions. An interactive tool is provided for prediction of substrate specificity. Interactive alignments for a suite of catalytic-core TyrA domains of differing specificity are also provided to facilitate phylogenetic analysis. tyrA membership in apparent operons (or supraoperons) was examined, and patterns of conserved synteny in relationship to organismal positions on the 16S rRNA tree were ascertained for members of the domain Bacteria. A number of aromatic-pathway genes (hisHb, aroF, aroQ) have fused with tyrA, and it must be more than coincidental that the free-standing counterparts of all of the latter fused genes exhibit a distinct trace of syntenic association.ConclusionWe propose that the ancestral TyrA dehydrogenase had broad specificity for both the cyclohexadienyl and pyridine nucleotide substrates. Indeed, TyrA proteins of this type persist today, but it is also common to find instances of narrowed substrate specificities, as well as of acquisition via gene fusion of additional catalytic domains or regulatory domains. In some clades a qualitative change associated with either narrowed substrate specificity or gene fusion has produced an evolutionary "jump" in the vertical genealogy of TyrA homologs. The evolutionary history of gene organizations that include tyrA can be deduced in genome assemblages of sufficiently close relatives, the most fruitful opportunities currently being in the Proteobacteria. The evolution of TyrA proteins within the broader context of how their regulation evolved and to what extent TyrA co-evolved with other genes as common members of aromatic-pathway regulons is now feasible as an emerging topic of ongoing inquiry.

Dataset Information

A core catalytic domain of the TyrA protein family: arogenate dehydrogenase from Synechocystis.

Publications

A core catalytic domain of the TyrA protein family: arogenate dehydrogenase from Synechocystis.

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