Structural and dynamics studies of the D54A mutant of human T cell leukemia virus-1 capsid protein.
The human T cell leukemia virus and the human immunodeficiency virus share a highly conserved, predominantly helical two-domain mature capsid (CA) protein structure with an N-terminal beta-hairpin. Despite overall structural similarity, differences exist in the backbone dynamic properties of the CA N-terminal domain. Since studies with other retroviruses suggest that the beta-hairpin is critical for formation of a CA-CA interface, we investigated the functional role of the human T cell leukemia virus beta-hairpin by disrupting the salt bridge between Pro(1) and Asp(54) that stabilizes the beta-hairpin. NMR (15)N relaxation data were used to characterize the backbone dynamics of the D54A mutant in the context of the N-terminal domains, compared with the wild-type counterpart. Moreover, the effect of the mutation on proteolytic processing and release of virus-like particles (VLPs) from human cells in culture was determined. Conformational and dynamic changes resulting from the mutation were detected by NMR spectroscopy. The mutation also altered the conformation of mature CA in cells and VLPs, as reflected by differential antibody recognition of the wild-type and mutated CA proteins. In contrast, the mutation did not detectably affect antibody recognition of the CA protein precursor or release of VLPs assembled by the precursor, consistent with the fact that the hairpin cannot form in the precursor molecule. The particle morphology and size were not detectably affected. The results indicate that the beta-hairpin contributes to the overall structure of the mature CA protein and suggest that differences in the backbone dynamics of the beta-hairpin contribute to mature CA structure, possibly introducing flexibility into interface formation during proteolytic maturation.
Bouamr
Fadila
F
Howard Hughes Medical Institute, New York, New York, USA.
Cornilescu
Claudia C
CC
Goff
Stephen P
SP
Tjandra
Nico
N
Carter
Carol A
CA
D036022:Capsid Proteins / Q000737:chemistry* / Q000235:genetics*; D002460:Cell Line; D003001:Cloning, Molecular; D017124:Conserved Sequence; D015368:Human T-lymphotropic virus 1 / Q000737:chemistry* / Q000235:genetics; D006801:Humans; D020125:Mutation, Missense; D019906:Nuclear Magnetic Resonance, Biomolecular; D018583:Pliability; D017433:Protein Structure, Secondary; D012996:Solutions
D036022:Capsid Proteins; D012996:Solutions