In vitro membrane integration of leader peptidase depends on the Sec machinery and anionic phospholipids and can occur post-translationally.
A cell-free system based on a lysate and membrane vesicles from Escherichia coli is used to study characteristics of the membrane integration reaction of the polytopic membrane protein leader peptidase (Lep). Integration into inverted inner membrane vesicles was detected by partial protection against externally added protease. Integration is most efficient when coupled to translation but can also occur post-translationally and depends on the action of the proteinaceous Sec machinery and availability of anionic phospholipids. Lep is the first example of a membrane protein without cleavable signal sequence which requires anionic lipids for integration in vitro.
van Klompenburg
W
W
Department of Biochemistry of Membranes, Centre for Biomembranes and Lipid Enzymology, Utrecht University, The Netherlands. w.klompenburg@biol.rug.nl
Ridder
A N
AN
van Raalte
A L
AL
Killian
A J
AJ
von Heijne
G
G
de Kruijff
B
B
D000251:Adenosine Triphosphatases / Q000276:immunology; D001425:Bacterial Outer Membrane Proteins / Q000378:metabolism; D001426:Bacterial Proteins / Q000276:immunology / Q000378:metabolism* / Q000502:physiology; D002474:Cell-Free System; D004926:Escherichia coli; D029968:Escherichia coli Proteins; D007425:Intracellular Membranes / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism*; D026901:Membrane Transport Proteins; D010743:Phospholipids / Q000502:physiology*; D011112:Polymyxin B / Q000494:pharmacology; D011498:Protein Precursors / Q000378:metabolism; D000069816:SEC Translocation Channels; D000081416:SecA Proteins; D012697:Serine Endopeptidases / Q000378:metabolism*
D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D008565:Membrane Proteins; D026901:Membrane Transport Proteins; D010743:Phospholipids; D011498:Protein Precursors; D000069816:SEC Translocation Channels; C057841:SecY protein, E coli; C057184:outer membrane protein A precursor (E coli); D012697:Serine Endopeptidases; C026515:type I signal peptidase; D000251:Adenosine Triphosphatases; D000081416:SecA Proteins; D011112:Polymyxin B