Conformational behaviour of a synthetic peptide of the C-terminus of villin that interacts with actin: an NMR, CD and stimulated annealing study.
The solution structure of a synthetic 22-amino acid peptide (P1) corresponding to the extreme C-terminal end and one of the F-actin binding sites of villin has been determined by 1H NMR and CD spectroscopy. The structure of this peptide was compared to that of a peptide in which lysine to glutamic acid substitutions were introduced at positions 17 and 19 (P11), abolishing F-actin binding. Both peptides are largely unstructured in aqueous solution. Changes observed in the NMR and CD spectra of both peptides are consistent with alpha-helix formation in trifluoroethanol/water mixtures. A set of 189 interproton distances derived from nuclear Overhauser enhancement (NOE) measurements, 17 phi-angle constraints obtained from 3JNH alpha coupling constants, as well as about 10 N ... O distance restraints deduced from amide proton exchange kinetics with deuterium, were used for the structure determination. The three-dimensional structure of P1 and P11 is characterized by two helical regions, one extending from residues 2 to 5 and a second covering residues 7 to 17. The central fragment, ranging from Leu-7 to Leu-15, is more stable. The C-terminal residues are less structured, particularly within peptide P11. The significance of these structural results is discussed in relation to the biological activity of villin.
Simenel
C
C
NMR Laboratory, Pasteur Institute, CNRS URA 1129, Paris, France.
Rose
T
T
Goethals
M
M
Vandekerckhove
J
J
Friederich
E
E
Louvard
D
D
Delepierre
M
M
D000199:Actins / Q000737:chemistry*; D000595:Amino Acid Sequence; D002135:Calcium-Binding Proteins / Q000737:chemistry*; D002264:Carboxylic Acids; D002352:Carrier Proteins / Q000737:chemistry*; D002942:Circular Dichroism; D003198:Computer Simulation; D009682:Magnetic Resonance Spectroscopy; D008840:Microfilament Proteins / Q000737:chemistry*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D013696:Temperature
D000199:Actins; D002135:Calcium-Binding Proteins; D002264:Carboxylic Acids; D002352:Carrier Proteins; D008840:Microfilament Proteins; C026615:villin