Tryptophan environment, secondary structure and thermal unfolding of the galactose-specific seed lectin from Dolichos lablab: fluorescence and circular dichroism spectroscopic studies.
Fluorescence and circular dichroism spectroscopic studies were carried out on the galactose-specific lectin from Dolichos lablab seeds (DLL-II). The microenvironment of the tryptophan residues in the lectin under native and denaturing conditions were investigated by quenching of the intrinsic fluorescence of the protein by a neutral quencher (acrylamide), an anionic quencher (iodide ion) and a cationic quencher (cesium ion). The results obtained indicate that the tryptophan residues of DLL-II are largely buried in the hydrophobic core of the protein matrix, with positively charged side chains residing close to at least some of the tryptophan residues under the experimental conditions. Analysis of the far UV CD spectrum of DLL-II revealed that the secondary structure of the lectin consists of 57% alpha-helix, 21% beta-sheet, 7% beta-turns and 15% unordered structures. Carbohydrate binding did not significantly alter the secondary and tertiary structures of the lectin. Thermal unfolding of DLL-II, investigated by monitoring CD signals, showed a sharp transition around 75 degrees C both in the far UV region (205 nm) and the near UV region (289 nm), which shifted to ca. 77-78 degrees C in the presence of 0.1 M methyl-beta-D-galactopyranoside, indicating that ligand binding leads to a moderate stabilization of the lectin structure.
Sultan
Nabil Ali Mohammed
NA
School of Chemistry, University of Hyderabad, Hyderabad-500 046, India.
Rao
Rameshwaram Nagender
RN
Nadimpalli
Siva Kumar
SK
Swamy
Musti J
MJ
D000373:Agglutinins / Q000737:chemistry; D002586:Cesium / Q000737:chemistry; D002942:Circular Dichroism; D031294:Dolichos / Q000378:metabolism*; D005690:Galactose / Q000737:chemistry; D007454:Iodides / Q000737:chemistry; D007477:Ions; D037102:Lectins / Q000737:chemistry*; D011489:Protein Denaturation; D017433:Protein Structure, Secondary; D012639:Seeds / Q000378:metabolism; D013050:Spectrometry, Fluorescence; D013696:Temperature; D014364:Tryptophan / Q000737:chemistry*
D000373:Agglutinins; D007454:Iodides; D007477:Ions; D037102:Lectins; D002586:Cesium; D014364:Tryptophan; D005690:Galactose