MOLECULAR REQUIREMENTS FOR THE INHIBITION OF THE TETRACYCLINE ANTIPORT PROTEIN AND THE EFFECT OF POTENT INHIBITORS ON THE GROWTH OF TETRACYCLINE-RESISTANT BACTERIA

Forty-seven compounds and tetracycline (Tc) structural analogues were tested for their ability to interfere with [H-3]Tc uptake in everted i nner membrane vesicles derived from Tc-resistant Escherichia coli D1-2 09, bearing the class B tetracycline resistance efflux protein (Tet pr otein). For effective inhibition of Tc uptake, the molecule had to hav e an intact ABCD tetracyclic carbon skeleton and a conjugated phenolic beta-diketone substructure at positions 10-12a with the subsequent de velopment of keto-enol tautomerization. Molecular variations at carbon positions 2, 4, 5, 6, 7, 8, and 9 did not decrease, and some increase d, the inhibitory activity as compared to that of Tc. Among these comp ounds, the highest inhibition of uptake occurred with certain position 6 and 13 derivatives of 5-hydroxytetracycline. In a group of 13-(prop ylthio) derivatives of 5-OH-Tc [13-propyl, 13-(3-chloropropyl), and 13 -(2-carboxyethyl)] there was a correlation between uptake inhibitory a ctivity and antibacterial activity. The 13-(3-chloropropyl) derivative , with the best efflux inhibitory activity, exhibited synergistic acti vity when tested in combination with doxycycline against Tc-resistant E, coli bearing the class A or B determinant, against Staphylococcus a ureus bearing class K, and against Enterococcus faecalis bearing the c lass L determinant.: The 13-propyl analogue also showed high transport blocking activity and showed synergistic antibacterial activity again st E. coli bearing the class A determinant and additive activity again st the other Tc-resistant bacteria. The synergistic antibacterial acti vity of these compounds was not shown by the 13-[(2-carboxyethyl)thio] homologue, whose efflux blocking activity was 70-fold less. These fin dings suggest that multiple sites on the Tc molecule are available for synthetic modification toward the development of an effective Tc bloc king agent. Such compounds, used alone or in combination with a standa rd tetracycline, show improved antibacterial activity.