Purification and characterization of an aminoglycoside inactivating enzyme from Staphylococcus epidermidis FK109 that nucleotidylates the 4'- and 4"-hydroxyl groups of the aminoglycoside antibiotics.

Abstract

The resistance to aminoglycoside antibiotics in S. epidermidis FK109, is mediated by an enzyme that catalyzes transfer of the nucleotide monophosphate moiety from the nucleotide triphosphates to the 4''-hydroxyl group or the 4"-hydroxyl group, that is in the equatorial plane of the aminoglycoside molecule. The enzyme, modifying the 2 sites, appears as a single and homogeneous entity in affinity chromatography, in chromatography on DEAE-Sepharose Cl-6B, in isoelectric focusing and in gel-filtration. It requires divalent cations, notably Mg++, and dithiothreitol for optimal adenylylation. It has a MW of 46,770 and an isoelectric point of 5.0. The ability of the enzyme modify the 2 hydroxyl groups of aminoglycoside molecules, enables it to have a spectrum of substrates that surpasses, in range, the substrate spectrum of all the aminoglycoside-modifying enzymes which were previously characterized.