Cloning and characterization of a eukaryotic pantothenate kinase gene (panK) from Aspergillus nidulans

Pantothenate kinase (PanK) is the key regulatory enzyme in the CoA biosynth etic pathway, The PanK gene from Escherichia coli (coaA) has been previousl y cloned and the enzyme biochemically characterized; highly related genes e xist in other prokaryotes. We isolated a PanK cDNA clone from the eukaryoti c fungus Aspergillus nidulans by functional complementation of a temperatur e-sensitive E. coli PanK mutant. The cDNA clone allowed the isolation of th e genomic clone and the characterization of the A. nidulans gene designated panK. The panK gene is located on chromosome (linkage group III), is inter rupted by three small introns, and is expressed constitutively. The amino a cid sequence of A, nidulans PanK (aPanK) predicted a subunit size of 46.9 k Da and bore little resemblance to its bacterial counterpart, whereas a high ly related protein was detected in the genome of Saccharomyces cerevisiae. In contrast to E. coli PanK (bPanK), which is regulated by CoA and to a les ser extent by its thioesters, aPanK activity was selectively and potently i nhibited by acetyl-CoA, Acetyl-CoA inhibition of aPanK was competitive with respect to ATP. Thus, the eukaryotic PanK has a distinct primary structure and unique regulatory properties that clearly distinguish it from its prok aryotic counterpart.