ABSTRACT
The human T cell receptor CD4 is a type I integral membrane glycoprotein that is involved in T cell activation and also acts as the primary coreceptor for human immunodeficiency viruses (HIV). Here the structure of a synthetic 38 amino acid peptide corresponding to the complete cytoplasmic domain of CD4 (CD4CYTO) has been investigated under a variety of solution conditions using a combination of circular dichroism and homonuclear two-dimensional 1H nuclear magnetic resonance spectroscopy. In the presence of the membrane mimetic 2,2,2-trifluoroethanol (TFE), a conformational change of CD4CYTO from a random coil to an alpha-helical structure was observed. In keeping with this, CD4CYTO has the potential to associate with membranes as demonstrated by binding studies of in vitro phosphorylated CD4CYTO with microsomal membranes. Both chemical shift and nuclear Overhauser enhancement data in 50% 2,2, 2-trifluoroethanol solution provide direct experimental evidence for the predominance of a short amphiphatic alpha-helix that is approximately 4 turns in length and extends from positions Arg-402 to Lys-417. The present data provide, for the first time, compelling experimental evidence that only a fraction of CD4CYTO has a propensity for adopting secondary structure under conditions that are assumed to exist at or near to the membrane surface and that this alpha-helical structure is located in the membrane-proximal region of CD4CYTO. The N-terminal residues, that link the alpha-helix to the transmembrane anchor of CD4, and a substantial C-terminal portion (14-18 residues) of CD4CYTO are unstructured under the solution conditions investigated. Correlation of our structural data with recent studies on the biological activity of CD4CYTO indicates that the alpha-helix is of crucial importance for the interaction of CD4 with Nef and Vpu in the process of HIV-mediated CD4 down-regulation.
