A methenyl tetrahydromethanopterin cyclohydrolase and a methenyl tetrahydrofolate cyclohydrolase in Methylobacterium extorquens AM1

Recently it was found that Methylobacterium extorquens AM1 contains both te trahydromethanopterin (H4MPT) and tetrahydrofolate (H4F) as carriers of C-1 units. In this paper we report that the aerobic methylotroph contains a me thenyl H4MPT cyclohydrolase (0.9 U.mg(-1) cell extract protein) and a methe nyl H4F cyclohydrolase (0.23 U.mg(-1)). Both enzymes, which were specific f or their substrates, were purified and characterized and the encoding genes identified via the N-terminal amino acid sequence. The purified methenyl H 4MPT cyclohydrolase with a specific activity of 630 U.mg(-1) (V-max = 1500 U.mg(-1); K-m = 30 mu M) was found to be composed of two identical subunits of molecular mass 33 kDa. Its sequence was approximate to 40% identical to that of methenyl H4MPT cyclohydrolases from methanogenic archaea. The meth enyl H4F cyclohydrolase with a specific activity of 100 U.mg(-1) (V-max = 3 30 U.mg(-1); K-m = 80 mu M) was found to be composed of two identical subun its of molecular mass 22 kDa. Its sequence was not similar to that of methe nyl H4MPT cyclohydrolases or to that of other methenyl H4F cyclohydrolases. Based on the specific activities in cell extract and from the growth prope rties of insertion mutants it is suggested that the methenyl H4MPT cyclohyd rolase might have a catabolic, and the methenyl-H4F cyclohydrolase an anabo lic function in the C-1-unit metabolism of M. extorquens AM1.