Calmodulin binding properties of peptide analogues and fragments of the calmodulin-binding domain of simian immunodeficiency virus transmembrane glycoprotein 41

The calcium-regulatory protein calmodulin (CaM) can bind with high affinity to a region in the cytoplasmic C-terminal tail of glycoprotein 41 of simia n immunodeficiency virus (SN). The amino acid sequence of this region is (1 )DLWETLRRGGRW(13)ILAIPRRIRQGLELT(28)L. In this work, we have used near- and far-uv CD, and fluorescence spectroscopy, to study the orientation of this peptide with respect to CaM. We have also studied biosynthetically carbon- 13 methyl-Met calmodulin by H-1,C-13 heteronuclear multiple quantum coheren ce NMR spectroscopy. Two Trp-substituted peptides, SIV-W3F and SIV-W12F, we re utilized in addition to the intact SIV peptide. Two half-peptides, SIV-N (residues 1-13) and SIV-C(residues 13-28) were also synthesized and studie d. The spectroscopic results obtained with the SIV-W3F and SIV-WI2F peptide s were generally consistent with those obtained for the native SIV peptide. Like the native peptide, these two analogues bind with an alpha -helical s tructure as shown by CD spectroscopy. Fluorescence intermolecular quenching studies suggested binding of Trp3 to the C-lobe of CaM. Our NMR results sh ow that SIV-N carl bind to both lobes of calcium-CaM, and that it strongly favors binding to the C-terminal hydrophobic region of CaM. The SN-C peptid e binds with relatively low affinity to both halves of the protein. These d ata reveal that the intact SIV peptide binds with its N-terminal region to the carboxy-terminal region of CaM, and this interaction initiates the bind ing of the peptide. This orientation is similar to that of most other CaM-b inding domains. (C) 2000 John Wiley & Sons, Inc.