A novel complete reconstitution system for membrane integration of the simplest membrane protein.
A complete reconstitution system for membrane integration of the simplest protein was developed by means of defined factors. A mutant version of Pf3 coat protein, 3L-Pf3 coat, requires neither signal recognition particle/Sec factors nor a membrane potential for its integration into the cytoplasmic membrane of Escherichia coli. Although 3L-Pf3 coat is spontaneously integrated into liposomes composed of phospholipids, diacylglycerol completely blocks such spontaneous integrations at a physiological level. Under the conditions where spontaneous integration does not occur, 3L-Pf3 coat integration was absolutely dependent on a novel integration-stimulating factor. Combination of the PURE system, an in vitro translation system composed of the purified factors involved in translation in E. coli, with liposomes containing the highly purified integration-stimulating factor revealed multiple cycles of 3L-Pf3 coat integration, achieving the complete reconstitution of membrane integration. Based on the function of the factor, we propose that the factor is named MPIase (Membrane Protein Integrase).
Nishiyama
Ken-ichi
K
Institute of Molecular and Cellular Biosciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-0032, Japan. nishiyam@iwate-u.ac.jp
Maeda
Masahide
M
Abe
Masato
M
Kanamori
Takashi
T
Shimamoto
Keiko
K
Kusumoto
Shoichi
S
Ueda
Takuya
T
Tokuda
Hajime
H
D036022:Capsid Proteins / Q000235:genetics / Q000378:metabolism; D002462:Cell Membrane / Q000737:chemistry / Q000378:metabolism*; D003546:Cysteine Endopeptidases / Q000378:metabolism; D004075:Diglycerides / Q000737:chemistry / Q000494:pharmacology; D004926:Escherichia coli / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000378:metabolism; D017912:Inovirus; D008081:Liposomes / Q000737:chemistry / Q000378:metabolism; D008565:Membrane Proteins / Q000737:chemistry / Q000378:metabolism*; D014176:Protein Biosynthesis; D000069816:SEC Translocation Channels
D036022:Capsid Proteins; D004075:Diglycerides; D029968:Escherichia coli Proteins; D008081:Liposomes; D008565:Membrane Proteins; D000069816:SEC Translocation Channels; D003546:Cysteine Endopeptidases; C532221:mepS protein, E coli