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).
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
Ken-ichi
K
Nishiyama
Masahide
M
Maeda
Masato
M
Abe
Takashi
T
Kanamori
Keiko
K
Shimamoto
Shoichi
S
Kusumoto
Takuya
T
Ueda
Hajime
H
Tokuda
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