The Neisseria gonorrhoeae pilus protein is one of the major antigenic determinants on the cell's surface. It is comprised of identical subunits of approximately 18 kd and plays a role in the infectivity and virulence of the organism. We have cloned the gene encoding a gonococcal pilus protein into Escherichia coli, and, using one of these clones as a probe in hybridization studies, we have shown that conversion of the pilus positive to pilus negative state in N. gonorrhoeae involves chromosomal rearrangement. Although the pilus protein is produced by E. coli, it does not appear to be assembled on the surface of the cell in native form.