Abstract
Bacillus vitellinus, a butirosin-producing organism, was shown to possess butirosin 3'-phosphotransferase catalyzing the phosphorylation of butirosin A into butirosin A 3'-phosphate.
The enzyme was purified about 1200-fold from the cell-free extract of the organism by ammonium sulfate fractionation, affinity chromatography on butirosin A-Sepharose 4 B and two gel filtrations on Sephadex G-100.
The molecular weight of the enzyme was estimated to be about 30, 000 by gel filtration. The pH optimum was between 6.7 and 8.8. Mg2+ was required for maximal activity and could be partially replaced by Co2+. ATP and GTP were effective phosphoryl donors. The enzyme catalyzed the phosphorylation of aminoglycoside antibiotics such as butirosin A, butirosin B, xylostasin, ribostamycin, neomycin, paromomycin, kanamycin A and kanamycin B. The Km values for butirosin A and ATP were 4.0×10-6M and 5.6×10-5M, respectively. The enzyme was strongly inhibited by p-chloromercuribenzoate, Ag+ and Hg2+, and was competitively inhibited by 3'-deoxybutirosin A.
