Membrane topology of NADPH-cytochrome P450 reductase on the endoplasmic reticulum.

Topology of the membrane-anchoring segment of mouse NADPH-cytochrome P450 reductase in the endoplasmic reticulum membrane was elucidated. An N-glycosylation site was generated in the amino-terminal hydrophilic sequence of the reductase, and the mutated protein was expressed in a cell-free system in the presence of microsomal vesicles. The in vitro synthesized reductase protein was integrated into the microsomal membrane and N-glycosylated depending on the presence of signal recognition particles. We conclude that the amino-terminal membrane-anchoring segment of the reductase is a type I signal-anchor sequence which shows amino-terminus-lumen and carboxy-terminus-cytoplasm topology.