Insertion and glycosylation of Pf3-derived membrane proteins in microsomes.
To get insight into the insertion mechanism of small newly synthesized single-spanning membrane proteins, Pf3 coat protein mutants were constructed with potential glycosylation sites in the N-terminus. Some of these proteins, when synthesized in vitro in the presence of microsomes, became efficiently glycosylated, proving that they insert into the membrane and translocate their N-terminus to the lumenal side. Such Pf3 constructs also insert efficiently into Escherichia coli vesicles and even in pure lipid vesicles, suggesting a common mechanism, which might be spontaneous. Glycosylation was sensitive to changes in the amino acid sequence of the N-terminus, suggesting that it depends on the structure of the protein and/or its positioning with respect to the lipid-water interface.
Ridder
Anja
A
Department of Biochemistry of Membranes, Centre for Biomembranes and Lipid Enzymology, Institute of Biomembranes, Utrecht University, Utrecht, The Netherlands. a.n.j.a.ridder@chem.uu.nl
Thissen
Laura
L
Killian
Antoinette
A
de Kruijff
Ben
B
D000595:Amino Acid Sequence; D002213:Capsid / Q000235:genetics / Q000378:metabolism*; D036022:Capsid Proteins; D006031:Glycosylation; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D008861:Microsomes / Q000378:metabolism*; D008969:Molecular Sequence Data; D009068:Movement; D009154:Mutation; D014176:Protein Biosynthesis; D011499:Protein Processing, Post-Translational; D021381:Protein Transport
D036022:Capsid Proteins; D008565:Membrane Proteins; C084332:major coat protein, Pseudomonas phage Pf3