An amphiphilic region in the cytoplasmic domain of KdpD is recognized by the signal recognition particle and targeted to the Escherichia coli membrane.
The sensor protein KdpD of Escherichia coli is composed of a large N-terminal hydrophilic region (aa 1-400), four transmembrane regions (aa 401-498) and a large hydrophilic region (aa 499-894) at the C-terminus. KdpD requires the signal recognition particle (SRP) for its targeting to the membrane. Deletions within KdpD show that the first 50 residues are required for SRP-driven membrane insertion. A fusion protein of the green fluorescent protein (GFP) with KdpD is found localized at the membrane only when SRP is present. The membrane targeting of GFP was not observed when the first 50 KdpD residues were deleted. A truncated mutant of KdpD containing only the first 25 amino acids fused to GFP lost its ability to specifically interact with SRP, whereas a specific interaction between SRP and the first 48 amino acids of KdpD fused to GFP was confirmed by pull-down experiments. Conclusively, a small amphiphilic region of 27 residues within the amino-terminal domain of KdpD (aa 22-48) is recognized by SRP and targets the protein to the membrane. This shows that membrane proteins with a large N-terminal region in the cytoplasm can be membrane-targeted early on to allow co-translational membrane insertion of their distant transmembrane regions.
Maier
Katja S
KS
Institute of Microbiology, University of Hohenheim, 70599 Stuttgart, Germany.
Hubich
Stefanie
S
Liebhart
Helga
H
Krauss
Susanne
S
Kuhn
Andreas
A
Facey
Sandra J
SJ
D000595:Amino Acid Sequence; D002462:Cell Membrane / Q000737:chemistry / Q000378:metabolism*; D004926:Escherichia coli / Q000737:chemistry / Q000166:cytology / Q000235:genetics / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D049452:Green Fluorescent Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D011494:Protein Kinases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D021382:Protein Sorting Signals; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017384:Sequence Deletion; D018271:Signal Recognition Particle / Q000378:metabolism*
D029968:Escherichia coli Proteins; D021382:Protein Sorting Signals; D011993:Recombinant Fusion Proteins; D018271:Signal Recognition Particle; D049452:Green Fluorescent Proteins; C489580:KdpD protein, E coli; D011494:Protein Kinases