Mutant strains of Escherichia coli male cells defective in Ca2+,Mg2+-dependent ATPase (unc) were constructed and tested for their ability to form a complex between sex pili and the filamentous phage fd under conditions where either the membrane potential or the cellular concentration of ATP was lowered. The uncoupler carbonyl cyanide m-chlorophenylhydrazone and the respiratory inhibitor cyanide, as well as valinomycin-K+ and colicin E1, all markedly diminished complex formation, indicating that the maintenance of a membrane potential, but probably not the pH gradient, is essential for the formation of the complex. Since complex formation with freshly centrifuged cells (which initially lacked sex pili) as well as with preincubated cells (in which pre-existing pili were available for complex formation) was inhibited by exposure to the inhibitors, energy seems to be required for both the reappearance (probably assembly) and the maintenance of sex pili on the cell surface. Brief exposure of freshly centrifuged cells to arsenate resulted in only partial inhibition of complex formation. However, marked inhibition of complex formation was observed following exposure to arsenate of preincubated cells possessing sex pili. This indicates that compounds such as ATP may also be required for maintenance of sex pili on the cell surface.