Conformational changes in concanavalin A associated with demetallization and alpha-methylmannose binding studied by Fourier transform infrared spectroscopy.
Infrared spectra of concanavalin A have been obtained both in the absence and in the presence of the metal ions, Mn2+ and Ca2+, and the saccharide, alpha-methylmannose. Second derivative calculations have been used to determine the frequencies of the different amide I and II components. In the demetallized protein dissolved in H2O buffer, absorptions in the amide I, II and III regions at 1695 and 1634, 1532 and 1237 cm-1, respectively, are assigned to beta-structure, while absorptions at 1563 and both 1318 and 1343 cm-1 are assigned to turns and bends. After deuterium exchange, the residual amide II maximum in the difference spectrum shifts from 1538 to 1563 cm-1, indicating that exchange is faster in the beta-structure than in the turns. In the presence of Mn2+ and Ca2+, the amide II band component at 1532 cm-1 shifts 4-6 cm-1 to higher wavenumbers, and the amide I band component at 1634 shifts 1 cm-1 in the same direction, both in H2O and 2H2O buffers, suggesting changes in the hydrogen-bonding network of a large portion of the protein, particularly in the beta-sheet regions. The addition of alpha-methylmannose increases the magnitude of exchange from 55% to above 90%. Comparison with existing X-ray crystallographic data has been made, and the usefulness of FT-IR to complement this technique is discussed.
Alvarez
J
J
Department of Biochemistry and Chemistry, Royal Free Hospital School of Medicine, University of London, U.K.
Haris
P I
PI
Lee
D C
DC
Chapman
D
D
D002118:Calcium; D003208:Concanavalin A / Q000032:analysis; D005583:Fourier Analysis; D008345:Manganese; D008759:Methylglycosides / Q000032:analysis; D008766:Methylmannosides / Q000032:analysis; D011487:Protein Conformation; D013055:Spectrophotometry, Infrared / Q000379:methods; D014961:X-Ray Diffraction
D008759:Methylglycosides; D008766:Methylmannosides; D003208:Concanavalin A; D008345:Manganese; C008466:methylmannoside; D002118:Calcium