Membrane Binding Mechanism of an RNA Virus-capping Enzyme

The RNA replication complex of Semliki Forest virus is bound to cytoplasmic membranes via the mRNA-capping enzyme Nsp1. Here we have studied the structure and liposome interactions of a synthetic peptide245GSTLYTESRKLLRSWHLPSV264corresponding to the membrane binding domain of Nsp1. The peptide inter...

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Bibliographic Details
Published in:	The Journal of biological chemistry Vol. 275; no. 48; pp. 37853 - 37859
Main Authors:	Lampio, Anja, Kilpeläinen, Ilkka, Pesonen, Saara, Karhi, Kimmo, Auvinen, Petri, Somerharju, Pentti, Kääriäinen, Leevi
Format:	Journal Article
Language:	English
Published:	United States Elsevier Inc 01-12-2000 American Society for Biochemistry and Molecular Biology
Subjects:	Amino Acid Sequence Cell Membrane - enzymology Membrane Fusion Models, Molecular Molecular Sequence Data NSP1 protein Protein Conformation RNA Caps Semliki Forest virus Semliki forest virus - enzymology Sequence Homology, Amino Acid Tryptophan - chemistry
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Summary:	The RNA replication complex of Semliki Forest virus is bound to cytoplasmic membranes via the mRNA-capping enzyme Nsp1. Here we have studied the structure and liposome interactions of a synthetic peptide245GSTLYTESRKLLRSWHLPSV264corresponding to the membrane binding domain of Nsp1. The peptide interacted with liposomes only if negatively charged lipids were present that induced a structural change in the peptide from a random coil to a partially α-helical conformation. NMR structure shows that the α-helix is amphipathic, the hydrophobic surface consisting of several leucines, a valine, and a tryptophan moiety (Trp-259). Fluorescence studies revealed that this tryptophan intercalates in the bilayer to the depth of the ninth and tenth carbons of lipid acyl chains. Mutation W259A altered the mode of bilayer association of the peptide and abolished its ability to compete for membrane association of intact Nsp1, demonstrating its crucial role in the membrane association and function of Nsp1.
Bibliography:	ObjectType-Article-2 SourceType-Scholarly Journals-1 ObjectType-Feature-1 content type line 23 ObjectType-Article-1 ObjectType-Feature-2
ISSN:	0021-9258 1083-351X
DOI:	10.1074/jbc.M004865200