Control SV40-transformed human fibroblasts can be readily adapted to growth on medium containing galactose as sole hexose source (galactose-MEH). However, most cells from a line of SV40-transformed skin fibroblasts from a patient with galactosemia (galactose-1-phosphate uridylyltransferase (GALT) deficiency) died in galactose-MEM. Surviving cells of this line either grew in completely sugar-free media or had acquired significant amounts of GALT activity. Two presumptive revertant cell lines with GALT activity were characterized in detail. The expression of GALT in these two lines was stable in nonselective conditions. Each had different reaction maximum velocities with respect to uridine diphosphoglucose (UDPg) concentration as compared to residual activity in the parental cell strain or control cells. Both appeared to demonstrate heat-inactivation profiles for GALT than differed from the parental cells or controls. UDPG concentration was found to significantly alter the thermostability of GALT. A competitive radioimmunoassay for GALT showed that these two lines had amounts of the GALT protein comparable to that of the parental cell strain or control cells. The electrophoretic mobility of GALT from the two presumptive revertants was found to differ from control cells. It was concluded that structural gene changes were probably responsible for the apparent reversion in these lines.