Biosynthesis of phosphatidylinositol-glycan (PI-G)-anchored membrane proteins in cell-free systems: PI-G is an obligatory cosubstrate for COOH-terminal processing of nascent proteins.
It is generally recognized that nascent proteins destined to be processed to a phosphatidylinositol-glycan (PI-G)-anchored membrane form contain a hydrophobic signal peptide at both their NH2 and COOH termini. In previous studies we showed that rough microsomal membranes (RM) prepared from CHO cells can carry out COOH-terminal processing. We have now investigated RM prepared from many additional cell types, including frog oocytes, B cells, and T cells, and found that all are competent with respect to COOH-terminal processing. Exceptions were certain mutant T cells that had been shown to be defective at various steps of PI-G anchor biosynthesis [Sugiyama, E., De Gasperi, R., Urakaze, M., Chang, H.-M., Thomas, L. J., Hyman, R., Warren, C. D. & Yeh, E. T. H. (1991) J. Biol. Chem. 266, 12119-12122]. In one such defective mutant, COOH-terminal processing activity of RM could be restored either by transfecting the intact cells with the gene for the deficient step in PI-G synthesis or by adding PI-G extracts to the RM in vitro. Cleavage of the COOH-terminal signal peptide in the RM is therefore dependent on the presence of intact PI-G incorporated into the mature protein.
Kodukula
K
K
Department of Neurosciences, Roche Institute of Molecular Biology, Nutley, NJ 07110.
Amthauer
R
R
Cines
D
D
Yeh
E T
ET
Brink
L
L
Thomas
L J
LJ
Udenfriend
S
S
D000217:Acyltransferases / Q000378:metabolism; D000469:Alkaline Phosphatase / Q000378:metabolism; D019881:Aminoacyltransferases; D000818:Animals; D016466:CHO Cells; D002474:Cell-Free System; D006224:Cricetinae; D004721:Endoplasmic Reticulum / Q000378:metabolism; D017261:Glycosylphosphatidylinositols; D006367:HeLa Cells; D006801:Humans; D008565:Membrane Proteins / Q000378:metabolism*; D010716:Phosphatidylinositols / Q000378:metabolism*; D010920:Placenta / Q000201:enzymology; D011134:Polysaccharides / Q000378:metabolism*; D011499:Protein Processing, Post-Translational
D017261:Glycosylphosphatidylinositols; D008565:Membrane Proteins; D010716:Phosphatidylinositols; D011134:Polysaccharides; D000217:Acyltransferases; D019881:Aminoacyltransferases; C023175:transamidases; D000469:Alkaline Phosphatase