The Escherichia coli htrB gene was originally discovered because its insertional inactivation led to an exquisitely temperature-sensitive phenotype in rich media, i.e. the ability to form colonies at temperatures below 32 degrees C, but not above 33 degrees C. The htrB gene has been sequenced. It can potentially code for two proteins, with Mr values of 35,407 Da and 8669 Da, that are encoded by overlapping, divergent open reading frames. Our data are consistent with the 35,407 Da protein being HtrB. Northern blot analysis clearly shows that the monocistronic htrB message is not under heat-shock regulation. We have also sequenced the flanking DNA and have discovered a new gene, designated orf39.9, located immediately adjacent to htrB, but divergently transcribed.