Expression and identification of the strA-strB gene pair from streptomycin-resistant Erwinia amylovora

Deletions in either of the genes in the strA-strB gene pair of Erwinia amylovora plasmid pEa34 resulted in a dramatic decrease in streptomycin resistance (SmR), but SmR was restored to high levels by complementation. When strA and strB were cloned separately on a lacIq/Ptac-based expression vector in Escherichia coli, only the protein encoded by strA was produced. When a strong Shine-Dalgarno sequence was introduced and the start codon was located outside the double-stranded region of the stable stem-loop, the protein encoded by strB was produced. Biochemical analysis of the respective phosphorylated products demonstrated that strA and strB encoded aminoglycoside-3"-phosphotransferase (APH(3")-Ib) and aminoglycoside-6-phosphotransferase (APH(6)-Id), respectively. These data suggest that the high-level SmR in bacteria containing strA and strB is due to the combined action of the two enzymes.