title: The N-terminal hydrophobic domain of P450c21 is required for membrane insertion and enzyme stability.
abstract: Microsomal cytochromes P-450 are known to be integrated into smooth endoplasmic reticulum through their hydrophobic sequences located at the N termini. The length requirement of the membrane insertion signal was determined by the generation of six plasmids encoding mutant P450c21 that lacked various portions of the N-terminal hydrophobic domains. When they were transcribed and translated in vitro in the presence of endoplasmic reticulum membranes, mutant protein lacking more than a third of the first hydrophobic domain gradually lost the ability to insert into the membrane and stayed mostly in the soluble fraction when the first N-terminal hydrophobic domain was removed. The steady-state amount of the truncated proteins was progressively reduced in parallel to the extent of their N-terminal deletions, due to their fast degradation. This process was accompanied by a decrease in the enzymatic activity. Therefore, the first hydrophobic domain of P450c21 not only serves as a membrane targeting and anchoring domain, but it is also important for the in vivo protein stability.
authors.affiliation: Institute of Molecular Biology, Academia Sinica, Taipei, Taiwan, Republic of China.
authors.forename: L C
authors.initials: LC
authors.lastname: Hsu
authors.forename: M C
authors.initials: MC
authors.lastname: Hu
authors.forename: H C
authors.initials: HC
authors.lastname: Cheng
authors.forename: J C
authors.initials: JC
authors.lastname: Lu
authors.forename: B C
authors.initials: BC
authors.lastname: Chung
mesh_terms: D000818:Animals; D001692:Biological Transport; D004285:Dogs; D004795:Enzyme Stability; D006801:Humans; D007425:Intracellular Membranes / Q000201:enzymology*; D008861:Microsomes / Q000201:enzymology; D009154:Mutation; D014176:Protein Biosynthesis; D017384:Sequence Deletion; D013255:Steroid 21-Hydroxylase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
chemical_list: D013255:Steroid 21-Hydroxylase