LASER RAMAN INVESTIGATION OF CONFORMATION OF HUMAN IMMUNOGLOBULIN-G

Laser Raman spectra of human immunoglobulin G in neutral solution, and in the lyophilized and alkaline-denatured states are presented. In the spectrum of the native protein, the amide III band appears at 1240 cm-1 and is assigned to the presence of .beta.-sheet structure. From its intensity, the .beta.-structure content was evaluated to be 37 .+-. 4%. This result is supported by the strong amide I'' band at 1667 cm-1 and by the presence in the spectra of 2 bands at 991 and 1078 cm-1, respectively assigned to the C-C and C-N skeletal stretching modes. The differences between the spectrum of the lyophilized powder and that of the solution show that the lyophilization process induces conformational changes that perturb the local environment of some of the tryptophan residues and alter the secondary structure of immunoglobulin G. The .beta.-structure appears to be more uniform and more abundant in solution. When the protein is denatured at pH 11, the amide III and amide I'' bands, which become weaker and broader, shift in frequency from 1240 to 1248 cm-1 and from 1667 to 1656 cm-1, respectively. These changes indicate a decrease in the amount of .beta.-structure and a transition toward a much more disordered conformation. During the denaturation, the intensities of many bands of the aromatic chromophores change, notably the tryptophan peaks at 879, 1359 and 1573 cm-1.