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Inducible overproduction of the Aspergillus nidulans pentafunctional AROM protein and the type-I and -II 3-dehydroquinases from Salmonella typhi and Mycobacterium tuberculosis.

ABSTRACT: The aroQ gene of Mycobacterium tuberculosis, encoding a type-II 3-dehydroquinase, and the aroD gene of Salmonella typhi, encoding a type-I 3-dehydroquinase, have been highly overexpressed in Escherichia coli using the powerful trc promoter contained within the expression vector pKK233-2. The M. tuberculosis type-II 3-dehydroquinase has been purified in bulk from overproducing strains of E. coli to greater than 95% homogeneity. The protein is extremely heat-stable, is active as a homododecamer and has the lowest reported Km value of any type-II 3-dehydroquinase. The pentafunctional aromA gene of Aspergillus nidulans has been overexpressed more than 120-fold in an A. nidulans aromA- qutB- double mutant from a truncated quinate-inducible qutE promoter, such that the AROM protein is visible as a significant fraction (approx. 6%) in cell-free crude extracts. The M. tuberculosis aroQ gene has been fused to the same truncated qutE promoter and shown to encode quinate-inducible 3-dehydroquinase activity that allows a qutE- mutant strain of A. nidulans to utilize quinate as sole carbon source.

SUBMITTER: Moore JD

PROVIDER: S-EPMC1133140 | biostudies-other | 1992 Oct

REPOSITORIES: biostudies-other

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Characterization of the 3-dehydroquinase domain of the pentafunctional AROM protein, and the quinate dehydrogenase from Aspergillus nidulans, and the overproduction of the type II 3-dehydroquinase from neurospora crassa.

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| S-EPMC1137716 | biostudies-other

Efficient independent activity of a monomeric, monofunctional dehydroquinate synthase derived from the N-terminus of the pentafunctional AROM protein of Aspergillus nidulans.

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