title: Metal cofactors of lysine-2,3-aminomutase.
abstract: Lysine-2,3-aminomutase from Clostridium SB4 contains iron and sulfide in equimolar amounts, as well as cobalt, zinc, and copper. The iron and sulfide apparently constitute an Fe-S cluster that is required as a cofactor of the enzyme. Although no B12 derivative can be detected, enzyme-bound cobalt is a cofactor; however, the zinc and copper bound to the enzyme do not appear to play a role in its catalytic activity. These conclusions are supported by the following facts reported in this paper. Purification of the enzyme under anaerobic conditions increases the iron and sulfide content. Lysine-2,3-aminomutase purified from cells grown in media supplemented with added CoCl2 contains higher levels of cobalt and correspondingly lower levels of zinc and copper relative to enzyme from cells grown in media not supplemented with cobalt. The specific activity of the purified enzyme increases with increasing iron and sulfide content, and it also increases with increasing cobalt and with decreasing zinc and copper content. The zinc and copper appear to occupy cobalt sites under conditions of insufficient cobalt in the growth medium, and they do not support the activity of the enzyme. The best preparations of lysine-2,3-aminomutase obtained to date exhibit a specific activity of approximately 23 units/mg of protein and contain about 12 g atoms of iron and of sulfide per mol of hexameric enzyme. These preparations also contain 3.5 g atoms of cobalt per mol, but even the best preparations contain small amounts of zinc and copper. The sum of cobalt, zinc, and copper in all preparations analyzed to date corresponds to 5.22 +/- 0.75 g atoms per mol of enzyme. An EPR spectrum of the enzyme as isolated reveals a signal corresponding to high spin Co(II) at temperatures below 20 K. The signal appears as a partially resolved 59Co octet centered at an apparent g value of 7. The 59Co hyperfine splitting (approximately 35 G) is prominent at 4.2 K. These findings show that lysine-2,3-aminomutase requires Fe-S clusters and cobalt as cofactors, in addition to the known requirement for pyridoxal 5'-phosphate and S-adenosylmethionine.
authors.affiliation: Institute for Enzyme Research, Graduate School, College of Agricultural and Life Sciences, University of Wisconsin, Madison 53705.
authors.forename: R M
authors.initials: RM
authors.lastname: Petrovich
authors.forename: F J
authors.initials: FJ
authors.lastname: Ruzicka
authors.forename: G H
authors.initials: GH
authors.lastname: Reed
authors.forename: P A
authors.initials: PA
authors.lastname: Frey
mesh_terms: D000591:Amino Acid Isomerases / Q000737:chemistry*; D003013:Clostridium / Q000201:enzymology*; D003035:Cobalt / Q000737:chemistry*; D003038:Cobamides / Q000032:analysis; D004578:Electron Spin Resonance Spectroscopy; D019751:Intramolecular Transferases; D007501:Iron / Q000737:chemistry*; D013440:Sulfides / Q000032:analysis
chemical_list: D003038:Cobamides; D013440:Sulfides; D003035:Cobalt; D007501:Iron; D000591:Amino Acid Isomerases; D019751:Intramolecular Transferases; C021518:lysine 2,3-aminomutase