Two Subsites on the Active Center of Pig Kidney Trehalase

Abstract

A kinetic analysis of the active site of pig kidney trehalase was made by examining two types of inhibitors that are monosaccharide analogs and cause a Competitive inhibition of the trehalase. Trehalase hydrolyzes trehalose (a-D-glyCopyranoSyl a-D-glucopyranoside) to give an equimolar mixture of a-Dglucose and, by inversion of configuration, P-D-glUCOse. 1,4-Dideoxy-l,4-imino-D-arabinito1 is considered to be a transition state (glucosyl cation) analog, while methyl P-D-glucoside, 1,5-dideoxy-1,5-iminO-Dglucitol (1 -deoxynojirimycin), fagomine, and 1-epivalidamine are considered to be analogs of the p-Dglucose that is derived by hydrolysis of trehalose. These glucosyl cation inhibitor and P-D-glucose analog inhibitors competed with each other at the same site on the active center of pig kidney trehalase and were therefore put together in one group (group A). Methyl a-D-mannoside and 1-deoxymannojirimycin were also competitive inhibitors of trehalase and competed with each other for the same site. However, an inhibitor i n group A did not compete with the methyl a-D-mannoside or 1,5-dideoxy-l ,.%imino-D-mannito1 (I -deoxymannojirimycin). Thus these latter two inhibitors were placed in group B. These results support the hypothesis that the active center of trehalase may comprise two subsites, one for catalysis and one for recognition, that act separately on each of the glucoses of the trehalose. The catalysis site requires the correct D-glucose configuration at carbons 2, 3, 4, and 5 or a good superimposition onto the glucosyl cation intermediate. The C2 equatorial OH group of a glucopyranosyl residue appears to be important for binding at the catalytic site since I-deoxynojiriniycin is more tightly bound by two orders of magnitude over its 2-deoxy derivative, fagomine. The P-D-glUCOSe and glucosyl cation analogs best fit

Synthesis, Inhibition Properties, and Theoretical Study of the New Nanomolar Trehalase Inhibitor 1‐Thiatrehazolin: Towards a Structural Understanding of Trehazolin Inhibition

“…[10,11] Trehalases are inverting glycosidases; this suggests the presence of a catalytic acid group, together with a nucleophilic water molecule, in the active site of the enzyme. [12] Kinetic studies performed with porcine kidney trehalase in the presence of two types of competitive inhibitors [13] support the earlier hypothesis [14] that the active center of the enzyme may comprise two subsites, one for catalysis and one for recognition, acting separately on each glucose unit of trehalose. This conclusion could probably be extended to other trehalases.…”

Synthesis of Novel Iminosugar‐Based Trehalase Inhibitors by Cross‐Metathesis Reactions

“…Final catalytic hydrogenation afforded compound 11. Due to the reported activity against trehalases of some iminofuranoses, such as compound 8 [11] (Figure 1), we envisaged the possibility to expand our set of potential inhibitors also to pseudodisaccharides obtained by CM of C-allyl-iminofuranoses. Two different allyl-iminofuranoses were synthesized and used for the CM reaction (Scheme 4), pos- The synthesis of α-C-allyl iminoarabinofuranosyl derivative 24 (Scheme 4A) proceeded by Grignard addition to nitrone 28, [19] as recently described.…”

“…Hence, a few iminosugar-based compounds (such as 4-8, Figure 1) have been also proposed as trehalase inhibitors. [5,11,12] [a] Department of Biotechnology and Biosciences,tected allyl C-iminoglycosides as the key step in homo-or heterodimerization reactions. The target products, obtained with the CM reaction, were fully hydrogenated by catalytic hydrogenolysis, and preliminary biological screening of the products as inhibitors of commercially available porcine trehalase was performed.…”

Synthesis and biological evaluation of nojirimycin- and pyrrolidine-based trehalase inhibitors

“…Furthermore, it was also reported that pyrrolidine derivatives (i.e., DAB-1, 9 , Fig. 2) [19] may act as trehalase inhibitors, in particular as competitive inhibitors with affinity to the catalytic site [19]. In general, it is well known that a key issue in the design of glycosidase inhibitors is specificity, for example, 1-deoxynojirimycin ( 7 ) is a glycosidase inhibitor in the low micromolar range, but despite its activity it lacks specificity.…”