The minimal structural requirement of concanavalin A that retains its functional aspects.
A systematic investigation of the effects of several commonly used detergents on the conformation and function of concanavalin A at pH 7 in solution form was made by using circular dichroism (CD), intrinsic fluorescence, 1-anilino 8-sulphonic acid (ANS) binding, dynamic light scattering (DLS) and sugar inhibition assay. In the presence of 6.0 mM sodium dodecyl sulphate (SDS), an anionic detergent, and 0.8 mM cetyl tri methyl ammonium bromide (CTAB), a cationic detergent, intermediate states of concanavalin A were obtained having a negative CD peaks at 222 and 208 nm respectively, a characteristic of alpha-helix. These states also retained tertiary contacts with altered tryptophan environment and high ANS binding (exposed hydrophobic area) which can be characterized as molten globule states. Concanavalin A in the presence of 5.0 mM 3-[(3-cholamidopropyl) dimethyl-ammonio]-1-propanesulphonate (CHAPS), a zwitterionic detergent, and 0.07 mM brij-35, a non-ionic detergent, also exists in intermediate states. These intermediates (molten globules) had high ANS binding with native-like secondary (inherent beta-sheet) and tertiary structure. The intermediate states were characterized further by means of dynamic light-scattering measurements and kinetic data. To study the possible functional requirement of the minimum structure, the intermediate states characterized in the presence of detergents were shown to retain the activity with polysaccharide (dextran). The pattern of activity observed was brij-35 > CHAPS > CTAB > SDS. The specific binding and activity of concanavalin A with ovalbumin was investigated as a function of time by turbidity measurements. Cationic and anionic detergents showed significant effects on the structure of concanavalin A as compared with zwitterionic and non-ionic detergents.
Ahmad
Ejaz
E
Interdisciplinary Biotechnology Unit, Department of Biochemistry, Aligarh Muslim University, Aligarh, India.
Naeem
Aabgeena
A
Javed
Saleem
S
Yadav
Savita
S
Khan
Rizwan Hasan
RH
D002236:Carbohydrate Conformation; D002942:Circular Dichroism; D003208:Concanavalin A / Q000737:chemistry* / Q000378:metabolism*; D003902:Detergents / Q000737:chemistry; D007700:Kinetics; D008027:Light; D012542:Scattering, Radiation; D013050:Spectrometry, Fluorescence
D003902:Detergents; D003208:Concanavalin A