The secondary structure of echistatin from 1H-NMR, circular-dichroism and Raman spectroscopy.
Detailed biophysical studies have been carried out on echistatin, a member of the disintegrin family of small, cysteine-rich, RGD-containing proteins, isolated from the venom of the saw-scaled viper Echis carinatus. Analysis of circular-dichroism spectra indicates that, at 20 degrees C, echistatin contains no alpha-helix but contains mostly beta-turns and beta-sheet. Two isobestic points are observed as the temperature is raised, the conformational changes associated with that observed between 40 degrees C and 72 degrees C being irreversible. Raman spectra also indicate considerable beta-turn and beta-sheet (20%) structure and an absence of alpha-helical structure. Three of the four disulphide bridges are shown to be in an all-gauche conformation, while the fourth adopts a trans-gauche-gauche conformation. The 1H-NMR spectrum of echistatin has been almost fully assigned. A single conformation was observed at 27 degrees C with the four proline residues adopting only the trans conformation. A large number of backbone amide protons were found to exchange slowly, but no segments of the backbone were found to be in either alpha-helical or beta-sheet conformation. A number of turns could be characterised. An irregular beta-hairpin contains the RGD sequence in a mobile loop at its tip. Two of the four disulphide cross-links have been identified from the NMR spectra. The data presented in this paper will serve to define the structure of echistatin more closely in subsequent studies.
Saudek
V
V
Marion Merrell Dow Research Institute, Strasbourg, France.
Atkinson
R A
RA
Lepage
P
P
Pelton
J T
JT
D000595:Amino Acid Sequence; D002942:Circular Dichroism; D036341:Intercellular Signaling Peptides and Proteins; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D009842:Oligopeptides / Q000737:chemistry; D010455:Peptides; D011487:Protein Conformation; D013059:Spectrum Analysis, Raman; D013696:Temperature; D014757:Viper Venoms / Q000737:chemistry*
D036341:Intercellular Signaling Peptides and Proteins; D009842:Oligopeptides; D010455:Peptides; D014757:Viper Venoms; C057986:echistatin; C047981:arginyl-glycyl-aspartic acid