CD4 is a type I transmembrane glycoprotein of 58 kDa. It has an extracellular domain of 370 amino acids, one transmembrane (TM) span, and a short cytoplasmic region of 40 residues. CD4 is involved in antigen recognition. CD4 also serves human immunodeficiency virus (HIV) as the major receptor for T-cell infection via interaction of its extracellular domain with HIV 1 envelope glycoprotein gp120. The cytoplasmic region of CD4 interacts with various cellular factors (e.g. kinases) and is one target of HIV-1 proteins VpU and Nef.
HIV-1 viral protein U (VpU) is an 81-residue transmembrane protein containing a short extracellular region, a TM helix, and a cytoplasmic domain. Interaction of CD4 and VpU in the ER membrane via their cytoplasmic and TM regions initiates proteasomal degradation of newly synthesized CD4, thus downregulating CD4 from the surface of infected cells.
We produced recombinant polypeptides. CD4tmcyt contains the TM and cytoplasmic regions of CD4, VpUcyt the cytoplasmic domain of VpU. CD4mut is a cysteine free mutant of CD4tmcyt. Structure and dynamics of these polypeptides in membrane mimicking micelles were characterized by NMR spectroscopy.
Schematic representation of CD4mut structure in a DPC micelle.
Structure of VpUcyt in presence of DPC micelles. Overlay of backbone bonds of 20 lowest energy structures and ribbon diagram of micelle-associated VpUcyt. The two serines shown in stick and ball form a phosphorylation motif.
We further studied the interaction of CD4mut with VpUcyt in liposomes and DPC micelles.
Solid state NMR experiments on CD4mut and full length VpU in liposomes were done in collaboration with the lab of Henrike Heise.