The bovine papillomavirus E6 gene can independently transform mouse C127 cells. To characterize E6 in greater detail, we created 16 site-directed mutations in E6, including substitution mutations in the cysteine codons of the four Cys-X-X-Cys motifs that are conserved in all papillomavirus E6 proteins. Proteins mutated in six of the seven cysteines tested, as well as those lacking the nonconserved C-terminus, were stable in transfected cells but were unable to induce morphological transformation, indicating that these amino acids play an important role in the function of E6.