2,3-di-O-tetradecyl-1-O-(beta-D-glucopyranosyl)-sn-glycerol is a substrate for human glucocerebrosidase.
Glucocerebrosidase, the lysosomal enzyme that is deficient in patients with Gaucher's disease, hydrolyses non-physiological aryl beta-D-glucosides and glucocerebroside, its substrate in vivo. We document that 2,3,-di-O-tetradecyl-1-O-(beta-D-glucopyranosyl)-sn-glycerol (2,3,-di-14:0-beta-Glc-DAG) inhibits human placental glucocerebrosidase activity in vitro (Ki 0.18 mM), and the nature of inhibition is typical of a mixed-type pattern. Furthermore, 2,3-di-14:0-beta-Glc-DAG was shown to be an excellent substrate for the lysosomal beta-glucosidase (Km 0.15 mM; Vmax. 19.8 units/mg) when compared with the natural substrate glucocerebroside (Km 0.080 mM; Vmax. 10.4 units/mg). The observations that (i) glucocerebrosidase-catalysed hydrolysis of 2,3-di-14:0-beta-Glc-DAG is inhibited by conduritol B epoxide and glucosylsphingosine, and (ii) spleen and brain extracts from patients with Gaucher's disease are unable to hydrolyse 2,3-di-14:O-beta-Glc-DAG demonstrate that the same active site on the enzyme is responsible for catalysing the hydrolysis of 4-methylumbelliferyl beta-D-glucopyranoside, glucocerebroside and 2,3-di-14:O-beta-Glc-DAG. With the aid of computer modelling we have established that the oxygen atoms in 2,3-DAG-Glc at the C-1, C-4*, C-5* (the ring oxygen in glucose) and C-2 positions correspond topologically to the oxygens at C-1, C-4* and C-5* and the nitrogen atom attached to C-2 respectively in glucocerebroside (* signifies a carbon atom in glucose); furthermore, all of the distances with respect to overlap of corresponding heteroatoms range from 0.02 A to 0.77 A (0.002-0.077 nm). A root-mean-square deviation of 0.31 A (0.031 nm) was obtained when the energy-minimized structures of 2,3-di-14:O-beta-Glc-DAG and glucocerebroside were compared using the latter four heteroatom co-ordinates.
Glew
R H
RH
Department of Biochemistry, School of Medicine, University of New Mexico, Albuquerque 87131.
Gopalan
V
V
Hubbell
C A
CA
Devraj
R V
RV
Lawson
R A
RA
Diven
W F
WF
Mannock
D A
DA
D001667:Binding, Competitive; D001921:Brain / Q000201:enzymology; D002151:Calorimetry; D003198:Computer Simulation; D005260:Female; D005776:Gaucher Disease / Q000201:enzymology; D005962:Glucosylceramidase / Q000378:metabolism*; D005963:Glucosylceramides / Q000378:metabolism; D006017:Glycolipids / Q000138:chemical synthesis / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D008968:Molecular Conformation; D010920:Placenta / Q000201:enzymology; D011247:Pregnancy; D012016:Reference Values; D013154:Spleen / Q000201:enzymology; D013379:Substrate Specificity; D001617:beta-Glucosidase / Q000378:metabolism
D005963:Glucosylceramides; D006017:Glycolipids; C051386:1,2-di-O-tetradecyl-3-O-(glucopyranosyl)glycerol; D001617:beta-Glucosidase; D005962:Glucosylceramidase