Positive charges on the translocating polypeptide chain arrest movement through the translocon.
Polypeptide chains synthesized by membrane-bound ribosomes are translocated through, and integrated into, the endoplasmic reticulum (ER) membrane by means of the protein translocation channel, the translocon. Positive charges on the nascent chain determine the orientation of the hydrophobic segment as it is inserted into the translocon and enhance the stop-translocation of translocating hydrophobic segments. Here we show that positive charges temporarily arrested ongoing polypeptide chain movement through the ER translocon by electrostatic interaction, even in the absence of a hydrophobic segment. The C-terminus of the polypeptide chain was elongated during the arrest, and then the full-length polypeptide chain moved through the translocon. The translocation-arrested polypeptide was not anchored to the membrane and the charges were on the cytoplasmic side of the membrane. The arrest effect was prevented by negatively charged residues inserted into the positive-charge cluster, and it was also suppressed by high salt conditions. We propose that positive charges are independent translocation regulators that are more active than previously believed.
Graduate School of Life Science, University of Hyogo, Kouto Ako-gun, Hyogo 678-1297,  Japan.
Hidenobu
H
Fujita
Marifu
M
Yamagishi
Yuichiro
Y
Kida
Masao
M
Sakaguchi
D000818:Animals; D003593:Cytoplasm / Q000737:chemistry; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D007425:Intracellular Membranes / Q000378:metabolism; D008565:Membrane Proteins / Q000032:analysis; D010455:Peptides / Q000737:chemistry / Q000378:metabolism; D021382:Protein Sorting Signals; D021381:Protein Transport; D051381:Rats; D012270:Ribosomes / Q000378:metabolism; D012965:Sodium Chloride / Q000737:chemistry
D008565:Membrane Proteins; D010455:Peptides; D021382:Protein Sorting Signals; D012965:Sodium Chloride