title: Biosynthesis of phosphatidylinositol-glycan (PI-G)-anchored membrane proteins in cell-free systems: cleavage of the nascent protein and addition of the PI-G moiety depend on the size of the COOH-terminal signal peptide.
abstract: Nascent translation products of PI-G-anchored membrane proteins contain both NH2- and COOH-terminal signal sequences of approximately 15-30 residues that are removed during processing. Removal of the latter occurs concomitant with the addition of the PI-G moiety to the newly formed COOH terminus. In human placental alkaline phosphatase (PLAP) the COOH-terminal signal peptide contains 29 residues. An engineered form of PLAP, miniPLAP 208, containing the same NH2- and COOH-terminal signal peptides as PLAP, was used as a substrate for cell-free processing. A comparison was made with mutants (delta 202, delta 197, delta 184, and delta 179) truncated at the COOH terminus. Intact preprominiPLAP 208 and truncated delta 202 were processed to yield the same mature product which, by size and distribution between Triton X-114 and water before and after treatment with inositol-specific phospholipases, indicates that it contained the PI-G moiety. Mutants that were further truncated at the COOH terminus, miniPLAPs delta 197, delta 184, and delta 179, were processed only at their NH2 termini. Those portions of the COOH-terminal sequence in miniPLAPs delta 197 and delta 1984 that extended beyond residue 179 were not removed during processing.
authors.affiliation: Department of Neurosciences, Roche Research Center, Nutley, NJ 07110.
authors.forename: K
authors.initials: K
authors.lastname: Kodukula
authors.forename: D
authors.initials: D
authors.lastname: Cines
authors.forename: R
authors.initials: R
authors.lastname: Amthauer
authors.forename: L
authors.initials: L
authors.lastname: Gerber
authors.forename: S
authors.initials: S
authors.lastname: Udenfriend
mesh_terms: D000469:Alkaline Phosphatase / Q000378:metabolism*; D000595:Amino Acid Sequence; D002474:Cell-Free System; D004252:DNA Mutational Analysis; D015870:Gene Expression; D006017:Glycolipids / Q000378:metabolism*; D017261:Glycosylphosphatidylinositols; D008562:Membrane Glycoproteins / Q000378:metabolism*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010716:Phosphatidylinositols / Q000378:metabolism*; D011257:Pregnancy Proteins / Q000378:metabolism*; D011499:Protein Processing, Post-Translational; D021382:Protein Sorting Signals / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics; D011994:Recombinant Proteins / Q000378:metabolism; D012995:Solubility; D013329:Structure-Activity Relationship
chemical_list: D006017:Glycolipids; D017261:Glycosylphosphatidylinositols; D008562:Membrane Glycoproteins; D010716:Phosphatidylinositols; D011257:Pregnancy Proteins; D021382:Protein Sorting Signals; D012333:RNA, Messenger; D011994:Recombinant Proteins; C101480:miniplacental alkaline phosphatase; D000469:Alkaline Phosphatase