Structural analysis of membrane-bound retrovirus capsid proteins

We have developed a system for analysis of histidine‐tagged (His‐tagged) retrovirus core (Gag) proteins, assembled in vitro on lipid monolayers consisting of egg phosphatidylcholine (PC) plus the novel lipid DHGN. DHGN was shown to chelate nickel by atomic absorption spectrometry, and DHGN‐containin...

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Bibliographic Details
Published in:	The EMBO journal Vol. 16; no. 6; pp. 1199 - 1213
Main Authors:	Barklis, Eric, McDermott, Jason, Wilkens, Stephan, Schabtach, Eric, Schmid, M.F., Fuller, Stephen, Karanjia, Sonya, Love, Zachary, Jones, Russell, Rui, Yuanjui, Zhao, Xiumin, Thompson, David
Format:	Journal Article
Language:	English
Published:	Chichester, UK John Wiley & Sons, Ltd 17-03-1997
Subjects:	AIDS/HIV Amino Acid Sequence Animals Capsid - chemistry Capsid - genetics Capsid - ultrastructure Chelating Agents COS Cells Crystallization electron microscopy Gene Products, gag - chemistry Gene Products, gag - genetics Gene Products, gag - ultrastructure image analysis lipid monolayer Lipids Membranes, Artificial Microscopy, Electron Molecular Sequence Data Molecular Structure Moloney murine leukemia virus - chemistry Moloney murine leukemia virus - genetics Moloney murine leukemia virus - ultrastructure Nickel Retroviridae Proteins - chemistry Retroviridae Proteins - genetics Retroviridae Proteins - ultrastructure retrovirus Transfection
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Summary:	We have developed a system for analysis of histidine‐tagged (His‐tagged) retrovirus core (Gag) proteins, assembled in vitro on lipid monolayers consisting of egg phosphatidylcholine (PC) plus the novel lipid DHGN. DHGN was shown to chelate nickel by atomic absorption spectrometry, and DHGN‐containing monolayers specifically bound gold conjugates of His‐tagged proteins. Using PC+DHGN monolayers, we examined membrane‐bound arrays of an N‐terminal His‐tagged Moloney murine leukemia virus (M‐MuLV) capsid (CA) protein, His‐MoCA, and in vivo studies suggest that in vitro‐derived His‐MoCA arrays reflect some of the Gag protein interactions which occur in assembling virus particles. The His‐MoCA proteins formed extensive two‐dimensional (2D) protein crystals, with reflections out to 9.5 Å resolution. The image‐analyzed 2D projection of His‐MoCA arrays revealed a distinct cage‐like network. The asymmetry of the individual building blocks of the network led to the formation of two types of hexamer rings, surrounding protein‐free cage holes. These results predict that Gag hexamers constitute a retrovirus core substructure, and that cage hole sizes define an exclusion limit for entry of retrovirus envelope proteins, or other plasma membrane proteins, into virus particles. We believe that the 2D crystallization method will permit the detailed analysis of retroviral Gag proteins and other His‐tagged proteins.
Bibliography:	istex:5C421663B29C149B6F2B37A69A5FC7AD98236BCB ArticleID:EMBJ7590112 ark:/67375/WNG-2J804L67-8 ObjectType-Article-2 SourceType-Scholarly Journals-1 ObjectType-Feature-1 content type line 23 ObjectType-Article-1 ObjectType-Feature-2
ISSN:	0261-4189 1460-2075 1460-2075
DOI:	10.1093/emboj/16.6.1199