Purification and characterization of a dual function 3-dehydroquinate dehydratase from Amycolatopsis methanolica.
Studies on hydroaromatic metabolism in the actinomycete Amycolatopsis methanolica revealed that the organism grows rapidly on quinate (but not on shikimate) as sole carbon- and energy source. Quinate is initially converted into the shikimate pathway intermediate 3-dehydroquinate by an inducible NAD(+)-dependent quinate/shikimate dehydrogenase. 3-Dehydroquinate dehydratase subsequently converts 3-dehydroquinate into 3-dehydroshikimate, which is used partly for the biosynthesis of aromatic amino acids, and is partly catabolized via protocatechuate and the beta-ketoadipate pathway. Enzyme studies and analysis of mutants clearly showed that the single 3-dehydroquinate dehydratase present in A. methanolica has a dual function, the first example of a 3-dehydroquinate dehydratase enzyme involved in both the catabolism of quinate and the biosynthesis of aromatic amino acids. This enzyme was purified over 1700-fold to homogeneity. Its further characterization indicated that it is a Type II 3-dehydroquinate dehydratase, a thermostable enzyme with a large oligomeric structure (native M(r) 135 x 10(3)) and a subunit M(r) of 12 x 10(3). Characterization of aromatic amino acid auxotrophic mutants of A. methanolica suggested that genes encoding 3-dehydroquinate synthase and 3-dehydroquinate dehydratase are genetically linked but their transcription results in the synthesis of two separate proteins.
Euverink
G J
GJ
Department of Microbiology, University of Groningen, Haren, The Netherlands.
Hessels
G I
GI
Vrijbloed
J W
JW
Coggins
J R
JR
Dijkhuizen
L
L
D000192:Actinomycetales / Q000201:enzymology*; D000595:Amino Acid Sequence; D006836:Hydro-Lyases / Q000302:isolation & purification*; D007700:Kinetics; D008969:Molecular Sequence Data; D008970:Molecular Weight; D016296:Mutagenesis; D011801:Quinic Acid / Q000031:analogs & derivatives / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D012765:Shikimic Acid / Q000378:metabolism
D011801:Quinic Acid; D012765:Shikimic Acid; D006836:Hydro-Lyases; C020000:3-dehydroquinate dehydratase