tailieunhanh - Báo cáo Y học: Artocarpus hirsuta lectin Differential modes of chemical and thermal denaturation

Unfolding, inactivation and dissociation of the lectin from Artocarpus hirsutaseedswere studiedbychemical (guanidine hydrochloride, GdnHCl) and thermal denaturation. Con-formational transitions were monitored by intrinsic ¯uor-escence and circular dichroism. The gradual red shift in the emissionmaxima of the native protein from335 to 356 nm, change intheellipticityat 218 nmandsimultaneousdecrease in the sugar binding activity were observed with increasing concentration of GdnHCl in the pH range between and . . | Eur. J. Biochem. 269 1413-1417 2002 FEBS 2002 Artocarpus hirsuta lectin Differential modes of chemical and thermal denaturation Sushama M. Gaikwad Madhura M. Gurjar and M. Islam Khan Division of Biochemical Sciences National Chemical Laboratory Pune India Unfolding inactivation and dissociation of the lectin from Artocarpus hirsuta seeds were studied by chemical guanidine hydrochloride GdnHCl and thermal denaturation. Conformational transitions were monitored by intrinsic fluorescence and circular dichroism. The gradual red shift in the emission maxima of the native protein from 335 to 356 nm change in the ellipticity at 218 nm and simultaneous decrease in the sugar binding activity were observed with increasing concentration of GdnHCl in the pH range between and . The unfolding and inactivation by GdnHCl were partially reversible. Gel filtration of the lectin in presence of 1-6 M GdnHCl showed that the protein dissociates reversibly into partially unfolded dimer and then irreversibly into unfolded inactive monomer. Thermal denaturation was irreversible. The lectin loses activity rapidly above 45 C. The exposure of hydrophobic patches distorted secondary structure and formation of insoluble aggregates of the thermally inactivated protein probably leads to the irreversible denaturation. Keywords Artocarpus lectin denaturation intrinsic fluorescence unfolding aggregation. Proteins that bind carbohydrates specifically and reversibly are termed as lectins. They occur ubiquitously in nature and have diverse role in plants animals and microbes. The recognition of carbohydrate moieties by lectins has important applications in a number of biological processes such as cell-cell interactions signal transduction and cell growth and differentiation 1 . a-Galactoside specific lectin present in the seeds of Artocarpus hirsuta 2-4 is a homotetrameric protein with molecular mass of 60 000 Da and high specificity for methyl a-D-galactopyranoside Me a-gal . The folding .

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