Kinetic study on conformational change in a single molecular species, beta3, of beta-conglycinin in an acidic ethanol solution.
The conformational change in a single molecular species, beta3, of beta-conglycinin in an acidic ethanol solution was kinetically studied by the stopped-flow technique, utilizing the intrinsic fluorescence of proteins and the fluorescence of 1-anilinonaphthalene-8-sulfonic acid (ANS) bound to the proteins. The time-course of the intrinsic fluorescence changes clearly showed the rate of conformational change below and above 25% ethanol to be quite different from each other. ANS could bind well to the protein in an ethanol concentration range of 15-25%. However, the rate of conformational change of the protein corresponding to that for ANS binding could not be obtained at less than 25% ethanol, while the rate of conformational change agreed well with that for ANS binding at more than 25% ethanol. In addition, the process showing the greatest and slowest ANS binding was not apparent in the denaturation of beta-conglycinin under the conditions employed. These results lead to the conclusions that the beta-conglycinin structure could be maintained in the mild molten globule-like denaturation state, and that various tertiary structural changes could take place without any significant effect on the high sensitivity of intrinsic fluorescence after the secondary structural changes.
Tsumura
Kazunobu
K
New Ingredients Research Institute, Tsukuba R&D Center, Fuji Oil Co. Ltd., 4-3 Kinunodai, Yawara, Tsukuba-gun, Ibaraki 300-2497, Japan. 830156@so.fujioil.co.jp
Kugimiya
Wataru
W
Kuwada
Masahiro
M
Shimura
Yuki
Y
Hasumi
Hideyo
H
D000817:Anilino Naphthalenesulfonates / Q000494:pharmacology; D052179:Antigens, Plant; D000431:Ethanol / Q000737:chemistry*; D005456:Fluorescent Dyes / Q000494:pharmacology; D005916:Globulins / Q000737:chemistry*; D007700:Kinetics; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D055314:Seed Storage Proteins; D012680:Sensitivity and Specificity; D030262:Soybean Proteins / Q000737:chemistry*; D013025:Glycine max / Q000378:metabolism; D013050:Spectrometry, Fluorescence; D013053:Spectrophotometry; D013997:Time Factors
D000817:Anilino Naphthalenesulfonates; D052179:Antigens, Plant; D005456:Fluorescent Dyes; D005916:Globulins; D055314:Seed Storage Proteins; D030262:Soybean Proteins; C013894:beta-conglycinin protein, Glycine max; D000431:Ethanol; C027132:1-anilino-8-naphthalenesulfonate