AN ABC-TRANSPORTER FROM STREPTOMYCES-LONGISPOROFLAVUS CONFERS RESISTANCE TO THE POLYETHER-IONOPHORE ANTIBIOTIC TETRONASIN

Streptomyces longisporoflavus produces the polyketide-polyether antibi otic, tetronasin, which acts as an ionophore and depolarizes the membr ane of bacteria sensitive to the drug. A genomic library of S. longisp oroflavus DNA was cloned in Streptomyces lividans and screened to iden tify tetronasin-resistance determinants. The inclusion of 0.2 M NaCl i n the growth medium with tetronasin markedly improved the sensitivity of the screen. Two different resistance determinants, designated tnrB (ptetR51) and tnrA (ptetR11) respectively, were identified. The determ inant tnrB (ptetR51) but not tnrA (ptetR11), also conferred resistance to tetronasin when cloned into Streptomyces albus. The tnrB determina nt was further localized, by subcloning, to a 2.8 kb Kpnl fragment. DN A sequence analysis of this insert revealed one incomplete and two com plete open reading frames (ORFs 1, 2 and 3). The deduced sequence of t he gene product of ORF2 (TnrBS) revealed significant similarity to the ATP-binding domains of the ABC (ATP binding cassette) superfamily of transport-related proteins. The adjacent gene, ORF3, is translationall y coupled to ORF2 and would encode a hydrophobic protein (TnrB3) with six transmembrane helices which probably constitutes the integral memb rane component of the transporter. The mechanism of tetronasin resista nce mediated by tnrB is probably an ATP-dependent efflux system.