Concerted influence of key amino acids on the lipid binding properties of a single-spanning membrane protein: NMR and mutational analysis.
Finding the combinations of key amino acids involved in the interaction network underlying the interfacial features of membrane proteins would contribute to a better understanding of their sequence-structure-function relationships and the role of anionic phospholipids. To further address these questions, we performed mutational analysis associated with NMR experiments on synthetic fragments of the single-spanning membrane protein PMP1 that exhibit binding specificity for phosphatidylserine (PS). The aromatic and glutamine residues of the helix part of the PMP1 cytoplasmic domain were mutated. (1)H NMR experiments were carried out using perdeuterated DPC micelles as a membrane-like environment, in the absence and presence of small amounts of either POPC or POPS lipids. From intermolecular NOEs and chemical shift data, specific and nonspecific aspects of peptide-phospholipid interactions were distinguished. The major finding of our study is to reveal the concerted influence of a tryptophan and a glutamine residue on the interfacial conformation and lipid binding specificity of the PMP1 cytoplasmic domain.
Mousson
F
F
Biophysique des Protéines et des Membranes, CEA DSV/DBCM and URA CNRS 2096, Centre d'Etudes de Saclay, 91191 Gif sur Yvette Cedex, France.
Beswick
V
V
Coïc
Y M
YM
Baleux
F
F
Huynh-Dinh
T
T
Sanson
A
A
Neumann
J M
JM
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000737:chemistry*; D002462:Cell Membrane / Q000737:chemistry*; D003593:Cytoplasm / Q000378:metabolism; D004252:DNA Mutational Analysis; D005973:Glutamine / Q000737:chemistry; D050356:Lipid Metabolism; D008055:Lipids / Q000737:chemistry; D009682:Magnetic Resonance Spectroscopy; D008823:Micelles; D008969:Molecular Sequence Data; D009154:Mutation; D010455:Peptides / Q000737:chemistry; D010718:Phosphatidylserines / Q000737:chemistry; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011522:Protons; D014364:Tryptophan / Q000737:chemistry
D000596:Amino Acids; D008055:Lipids; D008823:Micelles; D010455:Peptides; D010718:Phosphatidylserines; D011522:Protons; D005973:Glutamine; D014364:Tryptophan