Calmodulin disrupts the structure of the HIV-1 MA protein.
The MA protein from HIV-1 is a small, multifunctional protein responsible for regulating various stages of the viral replication cycle. To achieve its diverse tasks, MA interacts with host cell proteins and it has been reported that one of these is the ubiquitous calcium-sensing calmodulin (CaM), which is up-regulated upon HIV-1 infection. The nature of the CaM-MA interaction has been the subject of structural studies, using peptides based on the MA sequence, that have led to conflicting conclusions. The results presented here show that CaM binds intact MA with 1:1 stoichiometry in a Ca(2+)-dependent manner and that the complex adopts a highly extended conformation in solution as revealed by small-angle X-ray scattering. Alterations in tryptophan fluorescence suggest that the two buried tryptophans (W16 and W36) located in the first two alpha-helices of MA mediate the CaM interaction. Major chemical shift changes occur in the NMR spectrum of MA upon complex formation, whereas chemical shift changes in the CaM spectrum are quite modest and are assigned to residues within the normal target protein-binding hydrophobic clefts of CaM. The NMR data indicate that CaM binds MA via its N- and C-terminal lobes and induces a dramatic conformational change involving a significant loss of secondary and tertiary structure within MA. Circular dichroism experiments suggest that MA loses approximately 20% of its alpha-helical content upon CaM binding. Thus, CaM binding is expected to impact upon the accessibility of interaction sites within MA that are involved in its various functions.
Chow
John Y H
JY
School of Molecular Bioscience, University of Sydney, New South Wales 2006, Australia.
Jeffries
Cy M
CM
Kwan
Ann H
AH
Guss
J Mitchell
JM
Trewhella
Jill
J
D000595:Amino Acid Sequence; D002118:Calcium / Q000378:metabolism; D002147:Calmodulin / Q000737:chemistry* / Q000378:metabolism*; D002942:Circular Dichroism; D005453:Fluorescence; D015488:HIV Antigens / Q000737:chemistry* / Q000378:metabolism*; D015497:HIV-1 / Q000502:physiology*; D054884:Host-Pathogen Interactions; D006801:Humans; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011485:Protein Binding; D011487:Protein Conformation; D020836:Protein Structure, Quaternary; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D053838:Scattering, Small Angle; D054301:gag Gene Products, Human Immunodeficiency Virus / Q000737:chemistry* / Q000378:metabolism*
D002147:Calmodulin; D015488:HIV Antigens; D054301:gag Gene Products, Human Immunodeficiency Virus; C054858:p17 protein, Human Immunodeficiency Virus Type 1; D002118:Calcium