Compact acid-induced state of Clitoria ternatea agglutinin retains its biological activity.
The effects of pH on Clitoria ternatea agglutinin (CTA) were studied by spectroscopy, size-exclusion chromatography, and by measuring carbohydrate specificity. At pH 2.6, CTA lacks well-defined tertiary structure, as seen by fluorescence and near-UV CD spectra. Far-UV CD spectra show retention of 50% native-like secondary structure. The mean residue ellipticity at 217 nm plotted against pH showed a transition around pH 4.0 with loss of secondary structure leading to the formation of an acid-unfolded state. This state is relatively less denatured than the state induced by 6 M guanidine hydrochloride. With a further decrease in pH, this unfolded state regains ~75% secondary structure at pH 1.2, leading to the formation of the A-state with native-like near-UV CD spectral features. Enhanced 8-anilino-1-naphthalene-sulfonate binding was observed in A-state, indicating a "molten-globule" like conformation with exposed hydrophobic residues. Acrylamide quenching data exhibit reduced accessibility of quencher to tryptophan, suggesting a compact conformation at low pH. Size-exclusion chromatography shows the presence of a compact intermediate with hydrodynamic size corresponding to a monomer. Thermal denaturation of the native state was cooperative single-step transition and of the A-state was non-cooperative two-step transition. A-State regains 72% of the carbohydrate-binding activity.
Naeem
A
A
Interdisciplinary Biotechnology Unit, Aligarh Muslim University, Aligarh, 202002, India.
Saleemuddin
M
M
Khan
R H
RH
D000143:Acids / Q000494:pharmacology*; D000373:Agglutinins / Q000737:chemistry / Q000187:drug effects* / Q000502:physiology; D002942:Circular Dichroism; D031292:Clitoria / Q000737:chemistry*; D003064:Cnidarian Venoms / Q000737:chemistry*; D011487:Protein Conformation; D017510:Protein Folding
D000143:Acids; D000373:Agglutinins; D003064:Cnidarian Venoms