Studies on the Biosynthesis of Microsomal Membrane Proteins

• DOI: 10.1111/j.1432-1033.1982.tb05894.x
• Publication Date: 2005-03-03T21:51:51Z
• Authors: Yoshiie OKADA; Alan B. FREY; Thomas M. GUENTHNER; Franz OESCH; David D. SABATINI; Gert KREIBICH
• Keywords: Epoxide Hydrolases; Male; 0301 basic medicine; Immunodiffusion; Time Factors; Membrane Proteins; Rats, Inbred Strains; Rats; Electron Transport; 03 medical and health sciences; Cytochromes b5; Enzyme Induction; Phenobarbital; Protein Biosynthesis; Microsomes, Liver; Animals; Cytochromes; Rabbits; Cytochrome Reductases; Cytochrome-B(5) Reductase; NADPH-Ferrihemoprotein Reductase

The site of synthesis and mechanism of insertion into membranes of several microsomal polypeptides was studied using translation system programmed in vitro with polysomes or with mRNA extracted from free and membrane‐bound rat liver polysomes. Primary translation products of cytochrome b5, NADH: cytochrome b5 oxidoreductase, NADPH: cytochrome P‐450 oxidoreductase and epoxide hydrolase were isolated by specific immunoprecipitation and compared with the mature proteins. The following observations were made:

While cytochrome b5 and NADH: cytochrome b5 oxidoreductase are synthesized in free polysomes, NADPH: cytochrome P‐450 oxidoreductase and epoxide hydrolase are made in membrane‐bound polysomes. In all cases the radioactively labelled products synthesized in vitro comigrated with the purified native polypeptides. Since these proteins are not glycoproteins, it can be inferred that in vivo they do not undergo proteolytic cleavage during or after translation. Levels of translatable mRNA coding for NADPH: cytochrome P‐450 oxidoreductase or epoxide hydrolase were increased by phenobarbital or by various carcinogens respectively. A comparison of amino‐terminal segments of the native epoxide hydrolase and that synthesized in vitro showed that this protein retains an amino‐terminal polypeptide sequence which resembles the transient insertion signal found in presecretory proteins. The amino‐terminal region of NADPH: cytochrome P‐450 oxidoreductase contains 7 leucine residues in its first 15 amino acids and can, therefore, be presumed to be the hydrophobic portion which anchors this poly‐ peptide to the membrane.

Mechanisms which may account for the spatial disposition of the membrane proteins and for their selective accumulation in endoplasmic reticulum membranes are discussed.