External yeast beta-fructosidase. The role of tryptophyl residues in catalysis.
1. In native invertase at pH 4.6, 23 out of a total of 34 tryptophyl residues are "exposed" to oxidation with N-bromosuccinimide, the other residues being apparently shielded from the oxidant within the molecule. 2. Oxidation of 5-6 tryptophyl residues/molecule with N-bromosuccinimide is proportional to the complete inactivation of the enzyme, and appears to be specific for indole chromophore only. The ligand binding and fluorescence measurements indicate that the oxidation of native enzyme, up to 50% inhibition, apparently does not change the conformation and topography of enzymes surface. 3. Invertase is inhibited by diazonium-1-H-tetrazole. Since tyrosine residues can be excluded by nitration studies as catalytically unimportant, it appears that a mocification of a single histidyl residue/molecule with diazonium-1-H-tetrazole is sufficient to abolish the enzymic activity. However, the absence of inhibition with diethyl pyrocarbonate indicates that the inhibition with diazonium-1-H-tetrazole may be mediated through steric hindrance or other indirect effects. 4. The absence of inhibition with 2,4-dinitrophenylhydrazine, trinitro benzenesulfonic acid and 5,5'-dithiobis-(2-nitrobenzoate) indicates that the carbonyl groups of the carbohydrate moiety, free amino and -SH groups are not essential for activity.
D001665:Binding Sites; D001974:Bromosuccinimide / Q000494:pharmacology; D003979:Diazonium Compounds / Q000494:pharmacology; D004228:Dithionitrobenzoic Acid / Q000494:pharmacology; D005632:Fructose; D006026:Glycoside Hydrolases / Q000037:antagonists & inhibitors; D009578:Nitrobenzenes / Q000494:pharmacology; D010084:Oxidation-Reduction; D011485:Protein Binding; D011487:Protein Conformation; D012441:Saccharomyces cerevisiae / Q000201:enzymology*; D013050:Spectrometry, Fluorescence; D013393:Sucrase / Q000032:analysis*; D013777:Tetrazoles / Q000494:pharmacology; D014364:Tryptophan / Q000032:analysis
D003979:Diazonium Compounds; D009578:Nitrobenzenes; D013777:Tetrazoles; D005632:Fructose; D014364:Tryptophan; D004228:Dithionitrobenzoic Acid; D006026:Glycoside Hydrolases; D013393:Sucrase; D001974:Bromosuccinimide