Summary: The gene of , encoding a streptomycin 6-phosphotransferase (SPH), was sub-cloned in the pBR322-based expression vector pRK9 (which contains the promoter) with selection for expression of streptomycin resistance in Two hybrid plasmids, pCKL631 and pCKL711, were isolated which conferred resistance. Both contained a ~ 2 kbp fragment already suspected to include The properties of deletion derivatives of these plasmids were consistent with the presumed location deletion of a sequence including most of the promoter largely, but not quite completely, abolished the ability of the plasmid to confer streptomycin resistance, confirming that expression was indeed principally from the promoter. A polypeptide of ~ 34·5 kDa was present in minicells containing plasmids that conferred streptomycin resistance, but was absent when the plasmids contained deletions removing streptomycin resistance. Part of the fragment was sequenced and an open reading frame corresponding to identified. A computer-assisted comparison of the deduced SPH sequence with those of other antibiotic phosphotransferases suggested a common structure A-B-C-D-E, where B and D were conserved between all sequences compared while A, C and E divided between the streptomycin and hygromycin B phosphotransferases on one hand and kanamycin/neomycin ones on the other. A composite sequence database was searched for homologues to consensus matrices constructed from five approximately 12-residue subsequences within blocks B and D. For one subsequence, corresponding to the N-terminal portion of block D, those sequences from the database that yielded the highest homology scores comprised almost entirely either antibiotic phosphotransferases or eukaryotic protein kinases. Possible evolutionary implications of this homology, previously described by other groups, are discussed.


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