The vaccinia virus (VACV) protein A55 is a BTB/kelch protein with a broad-complex, tramtrack and bric-a-brac (BTB) domain in the N-terminal region and five kelch repeats in the C-terminal half. The BTB/kelch subgroup of the kelch superfamily of proteins has been associated with a wide variety of functions including regulation of the cytoskeleton. VACV contains three genes predicted to encode BTB/kelch proteins: A55R, F3L and C2L. The A55R gene product has been identified as an intracellular protein of 64 kDa that is expressed late in infection. A VACV strain lacking 93.6% of the A55R open reading frame (v Delta A55) was constructed and found to have an unaltered growth rate in vivo but a different plaque morphology and cytopathic effect, as well as reduced development of VACV-induced Ca2+-independent cell/extracellular matrix adhesion. In a murine intradermal model of VACV infection, a virus lacking the A55R gene induced larger lesions than wild-type and revertant control viruses.