Summary: The spectinomycin (Sp) resistance determinant from was cloned into using the plasmid vector pIJ699. A plasmid, pDGL15, with a 3.65 kb insert from conferring resistance to Sp was isolated. DNA sequence analysis of the 3651 bp DNA insert revealed four open reading frames (ORFs). The amino acid sequence deduced from one ORF (SpcN) showed a high degree of similarity to an aminoglycoside phosphotransferase (StrN) and from a second one (SpcR) to a regulatory protein (StrR) of the streptomycin biosynthesis gene cluster from The two other ORFs were incomplete and the deduced amino acid sequences showed similarities to an amidinotransferase encoded in the streptomycin biosynthesis gene cluster of and to the transposase of respectively. Expression of the gene in under the control of promoter conferred Sp resistance to the cells. An enzymic assay confirmed that the gene product of is an ATP-dependent aminoglycoside phosphotransferase which phosphoryiates Sp and actinamine, the aminocyclitol moiety of Sp.


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