Enzymatic Synthesis of Aminoglycoside Antibiotics: Novel Adenosylmethionine:2-Deoxystreptamine N-Methyltransferase Activities in Hygromycin B- and Spectinomycin-Producing Streptomyces spp. and Uses of the Methylated Products

Aminocyclitols structurally related to streptamine, a 1,3-diaminocyclitol, are common components of the RNA-binding aminoglycoside antibiotics. The respective aminocyclitol cores of hygromycin B and spectinomycin are N(3)-methyl-2-deoxy-d-streptamine and N(1),N(3)-dimethyl-2-epi-streptamine. Adenosyl[methyl-(14)C]methionine:2-deoxystreptamine N-methyltransferase activities were detected in extracts of early-stationary-phase mycelia of the hygromycin B producer Streptomyces hygroscopicus subsp. hygroscopicus ATCC 27438 and the spectinomycin producer Streptomyces flavopersicus ATCC 19756. Extracts of both strains methylated the N(1)- and N(3)-amino groups of 2-deoxystreptamine, streptamine, and 2-epi-streptamine; the N(1)-amino group of N(3)-methyl-2-deoxy-d-streptamine, and the N(3)-amino group of N(1)-ethyl-2-deoxy-d-streptamine, the semisynthetic aminocyclitol of netilmicin. The mono[(14)C]methyl derivatives of 2-deoxystreptamine, streptamine, and 2-epi-streptamine were excellent substrates for l-glutamine:aminocyclitol aminotransferase and thereby provided a sensitive assay for derepression of this key enzyme, a generic biosynthetic marker that we have shown to be the only enzyme common to the biosyntheses of all major aminoglycoside antibiotics. Other prospective uses for these methyl-labeled 2-deoxystreptamine analogs are also described.