title: Stability and kinetics of a bifunctional amylase/trypsin inhibitor.
abstract: The stability of the bifunctional amylase/trypsin inhibitor from ragi (Indian finger millet, Eleusine coracana) has been studied by methods of circular dichroism, UV absorption and intrinsic fluorescence. The inhibitor is stable in 8 M urea and 6 M guanidine-HCl. In 150 mM NaCl, thermal denaturation does not occur up to 90 degrees C. However, it is irreversibly denatured in 5 mM NaCl if heated over 73 degrees C. The acidic denaturation is reversible in both high and low salt conditions, but it shows different behavior below pH 1.65 under similar salt conditions. The helical content is about 2-4% in the pH range of 7-9 at which the inhibitor is active maximally. The NaCl concentration does not have a significant effect on the secondary structure elements. The beta-strand form does not show much variation under various conditions. Arg34-Leu35 is the reactive peptide bond in the trypsin-binding site. Trp and Tyr are involved in the binding with amylase. The bifunctional inhibitor represents the sum of individual inhibitors of trypsin and amylase.
authors.affiliation: Department of Biophysics, All India Institute of Medical Sciences, New Delhi.
authors.forename: S
authors.initials: S
authors.lastname: Alagiri
authors.forename: T P
authors.initials: TP
authors.lastname: Singh
mesh_terms: D000595:Amino Acid Sequence; D000681:Amylases / Q000037:antagonists & inhibitors*; D002942:Circular Dichroism; D019791:Guanidine; D006146:Guanidines; D006358:Hot Temperature; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008969:Molecular Sequence Data; D008897:Panicum / Q000737:chemistry*; D011489:Protein Denaturation; D017433:Protein Structure, Secondary; D013056:Spectrophotometry, Ultraviolet; D014357:Trypsin / Q000737:chemistry; D014361:Trypsin Inhibitors / Q000737:chemistry* / Q000302:isolation & purification; D014508:Urea
chemical_list: D006146:Guanidines; D014361:Trypsin Inhibitors; D014508:Urea; D000681:Amylases; D014357:Trypsin; D019791:Guanidine