Structure and function of PspA and Vipp1 N-terminal peptides: Insights into the membrane stress sensing and mitigation.
The phage shock protein (Psp) response maintains integrity of the inner membrane (IM) in response to extracytoplasmic stress conditions and is widely distributed amongst enterobacteria. Its central component PspA, a member of the IM30 peripheral membrane protein family, acts as a major effector of the system through its direct association with the IM. Under non-stress conditions PspA also negatively regulates its own expression via direct interaction with the AAA+ ATPase PspF. PspA has a counterpart in cyanobacteria called Vipp1, which is implicated in protection of the thylakoid membranes. PspA's and Vipp1's conserved N-terminal regions contain a putative amphipathic helix a (AHa) required for membrane binding. An adjacent amphipathic helix b (AHb) in PspA is required for imposing negative control upon PspF. Here, purified peptides derived from the putative AH regions of PspA and Vipp1 were used to directly probe their effector and regulatory functions. We observed direct membrane-binding of AHa derived peptides and an accompanying change in secondary structure from unstructured to alpha-helical establishing them as bona fide membrane-sensing AH's. The peptide-binding specificities and their effects on membrane stability depend on membrane anionic lipid content and stored curvature elastic stress, in agreement with full length PspA and Vipp1 protein functionalities. AHb of PspA inhibited the ATPase activity of PspF demonstrating its direct regulatory role. These findings provide new insight into the membrane binding and function of PspA and Vipp1 and establish that synthetic peptides can be used to probe the structure-function of the IM30 protein family.
Department of Life Sciences, Imperial College London, London SW7 2AZ, UK; Department of Chemistry and Institute of Chemical Biology, Imperial College London, London SW7 2AZ, UK.
Christopher
C
McDonald
Department of Life Sciences, Imperial College London, London SW7 2AZ, UK.
Goran
G
Jovanovic
Institute of Structural and Molecular Biology, Birkbeck College, University of London, London, WC1E 7HX, UK.
B A
BA
Wallace
Department of Chemistry and Institute of Chemical Biology, Imperial College London, London SW7 2AZ, UK.
Oscar
O
Ces
Department of Life Sciences, Imperial College London, London SW7 2AZ, UK. Electronic address: m.buck@imperial.ac.uk.
Martin
M
Buck
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry* / Q000378:metabolism; D002462:Cell Membrane / Q000737:chemistry / Q000378:metabolism; D004926:Escherichia coli / Q000737:chemistry / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000378:metabolism; D006360:Heat-Shock Proteins / Q000737:chemistry* / Q000378:metabolism; D008565:Membrane Proteins / Q000737:chemistry* / Q000378:metabolism; D010455:Peptides / Q000138:chemical synthesis / Q000737:chemistry*; D010713:Phosphatidylcholines / Q000737:chemistry* / Q000378:metabolism; D010715:Phosphatidylglycerols / Q000737:chemistry* / Q000378:metabolism; D011485:Protein Binding; D017433:Protein Structure, Secondary; D013312:Stress, Physiological; D046939:Synechocystis / Q000737:chemistry / Q000378:metabolism; D015534:Trans-Activators / Q000737:chemistry* / Q000378:metabolism; D053835:Unilamellar Liposomes / Q000737:chemistry / Q000378:metabolism
D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D006360:Heat-Shock Proteins; D008565:Membrane Proteins; D010455:Peptides; D010713:Phosphatidylcholines; D010715:Phosphatidylglycerols; C098954:PspF protein, E coli; D015534:Trans-Activators; D053835:Unilamellar Liposomes; C424449:VIPP1 protein, Synechocystis; C069816:phage shock protein, Bacteria; C051388:1,2-dioleoyl-sn-glycero-3-phosphoglycerol; C017251:1,2-oleoylphosphatidylcholine
Amphipathic helix conformation; Anionic lipids; Membrane structure; Peripheral membrane protein; Stored curvature elastic stress