Conformational features of the full-length HIV and SIV Nef proteins determined by mass spectrometry.
The Nef protein from human or simian immunodeficiency virus enhances viral replication, downregulates immune cell receptors, and activates multiple host cell signaling pathways. Conformational information about full-length Nef has been difficult to obtain as the full-length protein is not readily amenable to NMR or X-ray crystallography due to aggregation at high concentrations. As an alternative, full-length HIV and SIV Nef were probed with hydrogen exchange mass spectrometry, a method compatible with the low concentration requirements of Nef. The results showed that HIV Nef contains a solvent-protected core, as previously demonstrated with both NMR and X-ray crystallography. SIV Nef, for which there is no structural information, had a similar protected core, although it was more flexible and dynamic than its HIV counterpart. Many of the regions outside the core in both SIV and HIV Nef were highly solvent exposed. However, limited protection from exchange was observed in both N- and C-terminal regions, suggesting the presence of structured elements. Protection from exchange was also observed in a large loop emanating from the core that was deleted for NMR and X-ray analysis. These data show that while the majority of Nef was highly solvent exposed, regions outside the core may have structural attributes which may contribute to Nef functions known to map to these regions.
Hochrein
James M
JM
Department of Chemistry, University of New Mexico, Albuquerque, New Mexico 87131, USA.
Wales
Thomas E
TE
Lerner
Edwina C
EC
Schiavone
Anthony P
AP
Smithgall
Thomas E
TE
Engen
John R
JR
D000595:Amino Acid Sequence; D041961:Deuterium Exchange Measurement; D015693:Gene Products, nef / Q000737:chemistry*; D006678:HIV / Q000737:chemistry; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D011487:Protein Conformation; D016415:Sequence Alignment; D054334:Viral Regulatory and Accessory Proteins / Q000737:chemistry*; D054311:nef Gene Products, Human Immunodeficiency Virus
D015693:Gene Products, nef; C429284:NEF protein, SIV; D054334:Viral Regulatory and Accessory Proteins; D054311:nef Gene Products, Human Immunodeficiency Virus