Attempted affinity-labelling of beta-D-galactosidase from Escherichia coli with 2,6:3,4-dianhydro-1-deoxy-D-talo-hept-1-enitol.
The epoxides methyl 2,3-anhydro-beta-D-talopyranoside (1) and 2,6:3,4-dianhydro-1-deoxy-D-talo-hept-1-enitol (2), both prepared by improved methods, and 2,6:3,4-dianhydro-1-deoxy-D-gulo-hept-1-enitol (3) were applied as potential reagents for the affinity labelling of E. coli beta-D-galactosidase. Compounds 1 and 3 are ineffective as labelling reagents, whereas compound 2 irreversibly inhibits the enzyme activity. Deactivation is complete only when high concentrations (0.5M) of the inhibitor are applied over a relatively long period of time (24 h). Saturation kinetics cannot be observed. Nevertheless, the competitive inhibitor isopropyl 1-thio-beta-D-galactopyranoside protects the enzyme from irreversible deactivation by 2, indicating that the latter also reacts with the active site. Treatment of beta-D-galactosidase with 2,6:3,4-dianhydro-1-deoxy-D-talo-[4-3H]hept-1-enitol under conditions that effect deactivation of the enzyme to only a minor extent causes labelling of the protein at the molar ratio of 48:1. Specific, radioaffinity labelling of the active site of the enzyme cannot be thus achieved.
D000345:Affinity Labels / Q000494:pharmacology*; D004926:Escherichia coli / Q000201:enzymology*; D005696:Galactosidases / Q000378:metabolism*; D007700:Kinetics; D013329:Structure-Activity Relationship; D013402:Sugar Alcohols / Q000138:chemical synthesis / Q000494:pharmacology*; D001616:beta-Galactosidase / Q000378:metabolism*
D000345:Affinity Labels; D013402:Sugar Alcohols; C036066:2,6-3,4-dianhydro-1-deoxyhept-1-enitol; D005696:Galactosidases; D001616:beta-Galactosidase