PURIFICATION OF A NOVEL COENZYME F-420-DEPENDENT GLUCOSE-6-PHOSPHATE-DEHYDROGENASE FROM MYCOBACTERIUM-SMEGMATIS

A variety of Mycobacterium species contained the 5-deazaflavin coenzym e known as F-420. Mycobacterium smegmatis was found to have a glucose- 6-phosphate dehydrogenase that was dependent on F-420 as an electron a cceptor and which did not utilize NAD or NADP. The enzyme was purified by ammonium sulfate fractionation, phenyl-Sepharose column chromatogr aphy, F-420-ether-linked aminohexyl-Sepharose 4B affinity chromatograp hy, and quaternary aminoethyl-Sephadex column chromatography, and the sequence of the first 26 N-terminal amino acids has been determined. T he response of enzyme activity to a range of pHs revealed a two-peak p attern, with maxima at pH 5.5 and 8.0. The apparent K-m values for F-4 20 and glucose-6-phosphate were, respectively, 0.004 and 1.6 mM. The a pparent native and subunit molecular masses were 78,000 and similar to 40,000 Da, respectively.