J. Bacteriol., 05 1996, 2861-2866, Vol 178, No. 10
Copyright © 1996, American Society for Microbiology

Purification of a novel coenzyme F420-dependent glucose-6-phosphate dehydrogenase from Mycobacterium smegmatis

E Purwantini and L Daniels
Department of Microbiology, University of Iowa, Iowa City 52242, USA.

A variety of Mycobacterium species contained the 5-deazaflavin coenzyme known as F420. Mycobacterium smegmatis was found to have a glucose-6- phosphate dehydrogenase that was dependent on F420 as an electron acceptor and which did not utilize NAD or NADP. The enzyme was purified by ammonium sulfate fractionation, phenyl-Sepharose column chromatography, F420-ether-linked aminohexyl-Sepharose 4B affinity chromatography, and quaternary aminoethyl-Sephadex column chromatography, and the sequence of the first 26 N-terminal amino acids has been determined. The response of enzyme activity to a range of pHs revealed a two-peak pattern, with maxima at pH 5.5 and 8.0. The apparent Km values for F420 and glucose-6-phosphate were, respectively, 0.004 and 1.6 mM. The apparent native and subunit molecular masses were 78,000 and approximately 40,000 Da, respectively.

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