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Journal of Virology, July 2002, p. 6900-6908, Vol. 76, No. 14
0022-538X/02/$04.00+0     DOI: 10.1128/JVI.76.14.6900-6908.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

Kinetic Analysis of the Role of Intersubunit Interactions in Human Immunodeficiency Virus Type 1 Capsid Protein Assembly In Vitro

Jason Lanman,1 Jennifer Sexton,1 Michael Sakalian,2 and Peter E. Prevelige, Jr.1*

Department of Microbiology, University of Alabama at Birmingham, Birmingham, Alabama 35294,1 Department of Microbiology and Immunology, University of Oklahoma, Oklahoma City, Oklahoma 731902

Received 30 January 2002/ Accepted 8 April 2002

The human immunodeficiency virus type 1 (HIV-1) capsid protein (CA) plays a crucial role in both assembly and maturation of the virion. Numerous recent studies have focused on either the soluble form of CA or the polymer end product of in vitro CA assembly. The CA polymer, in particular, has been used to study CA-CA interactions because it is a good model for the CA interactions within the virion core. However, analysis of the process of in vitro CA assembly can yield valuable insights into CA-CA interactions and the mechanism of core assembly. We describe here a method for the analysis of CA assembly kinetics wherein the progress of assembly is monitored by using turbidity. At pH 7.0 the addition of either of the isolated CA domains (i.e., the N or the C domain) to an assembly reaction caused a decrease in the assembly rate by competing for binding to the full-length CA protein. At pH 8.0 the addition of the isolated C domain had a similar inhibitory affect on CA assembly. However, at pH 8.0 the isolated N domain had no affect on the rate of CA assembly but, when mixed with the C domain, it alleviated the C-domain inhibition. These data provide biochemical evidence for a pH-sensitive homotypic N-domain interaction, as well as for an N- and C-domain interaction.

* Corresponding author. Mailing address: Department of Microbiology, University of Alabama at Birmingham, BBRB 416, 845 19th St. S., Birmingham, AL 35294-2170. Phone: (205) 975-5327. Fax: (205) 975-5479. E-mail: prevelig{at}uab.edu.

Journal of Virology, July 2002, p. 6900-6908, Vol. 76, No. 14
0022-538X/02/$04.00+0     DOI: 10.1128/JVI.76.14.6900-6908.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.



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