UDP-glucuronosyltransferases (UGTs) are membrane-bound glycoproteins that are resident in the endoplasmic reticulum with a type I topology. The roles of the membrane-spanning and membrane-proximal cytoplasmic domains in UGT activity were investigated. Site-directed and deletional mutagenesis techniques were used to generate truncated forms of the enzyme, forms with altered residues, or forms with heterologous tails appended to the carboxyl terminus. The presence of the transmembrane domain was a critical requirement for UGT activity whereas the cytoplasmic domain seemed to be a modulator of activity but was not essential. Truncation of the protein did not appear to lead to scavenging and degradation, although appending long heterologous tails to the cytoplasmic domain did seem to trigger proteolysis. Analysis of enzyme kinetic parameters and enzyme latency allowed us to discount substrate binding or substrate transport defects as the cause of ameliorated UGT activity in the mutants.