PubMed1983
Values of kinetic inhibition constants representing dissociation constants of inhibitors interacting with the free form of beta-galactosidase ('galactose" site) were determined for a large number of monosaccharides and alcohols. The studies showed that when hydroxyl groups were present at positions 3, 4, and 6 and were in the same orientation as in galactose, binding to the 'galactose" site was good. Alterations in the position of or a lack of a hydroxyl at any of these three positions decreased binding dramatically. The overall binding specificity thus depends to a large extent on these three positions. Positions 3 and 4 were critical. A misorientation at either position eliminated most of the binding. Position 6 was a little more independent. The absence of a hydroxyl at that position did not totally eliminate the binding. The position 2 hydroxyl did not seem to be important for binding as its absence or orientation had only a small effect on the binding capacity. Studies regarding position 5 were inconclusive but since a minimum of four hydroxyl groups seemed important for binding and since positions 3, 4, and 6 are important the 5 position may also be of significance. The work also showed that p-nitrophenyl-beta-D-glucopyranoside, p-nitrophenyl-beta-L-arabinofuranoside, p-nitrophenyl-beta-D-xylopyranoside, cellobiose, and gentiobiose were not substrates in spite of the fact that the nitrophenyl groups made some of these bind quite well. Thus, not only are the presence and configuration of the hydroxyls at positions 3 and 4 important for binding, they are also important in catalysis. p-Nitrophenyl-beta-D-fucopyranoside was a substrate indicating that the 6 position hydroxyl is not as important for production binding as are the hydroxyls at positions 3 and 4.