Mechanism of inhibition of beta-hexosaminidase B from human liver by short- and medium-chain monocarboxylic acids.
Short- and medium-chain monocarboxylic acids showed an inhibitory effect on enzymatic activity of beta-hexosaminidase B (Hex B) when 4-methylumbelliferyl-2-acetamido-2-deoxyglucopyranoside (MU-GlcNAc) was used as substrate: 1. Two groups were distinguished according to the chain length of the monocarboxylic acids: the first was only constituted by acetic acid (C2) whereas the second group exhibited a broader chain length specificity for medium-chain monocarboxylic acids (between C6 and C9). 2. Both groups were reversible competitive inhibitors (Km = 0.52 +/- 0.15 mM; KiC2 = 21.4 +/- 3.0 mM; KiC7 = 3.4 +/- 0.5 mM). Competition experiments between C2 and C7 (as representent of medium-chain monocarboxylic acids group) demonstrated that these inhibitors were bound to different subsites. 3. Competition experiments between C2 and 2-acetamido-2-deoxy-D-galactonolactone (GalNAcLone) (a competitive inhibitor of lysosomal hexosaminidases) demonstrated that these two inhibitors were mutually exclusive, i.e. they were probably bound at the same subsite. This feature and the structural analogy of C2 with the acetyl residue of GalNAcLone (and of the saccharidic part of the substrate) suggested that C2 bound to the substrate site where the N-acetyl residue of the beta-N-acetyl hexosaminide was positioned. 4. The inhibitory effect of medium-chain monocarboxylic acids (C6 and C9) was dependent on their physical state. Below the critical micellar concentration (CMC), detected by a dye spectral shift method, no significant inhibition was detected, but as extensively reported using C7, an obvious inhibitory effect occurred at concentrations higher than CMC.(ABSTRACT TRUNCATED AT 250 WORDS)
Hardy
M
M
Laboratoire de Biochimie et INSERM Unité 101, Faculté de Médecine, Toulouse, France.
Salvayre
R
R
Maret
A
A
Douste-Blazy
L
L
D001665:Binding Sites; D001667:Binding, Competitive; D002264:Carboxylic Acids / Q000494:pharmacology*; D054819:Hexosaminidase B; D006801:Humans; D007700:Kinetics; D008099:Liver / Q000201:enzymology*; D013329:Structure-Activity Relationship; D001619:beta-N-Acetylhexosaminidases / Q000037:antagonists & inhibitors*
D002264:Carboxylic Acids; D054819:Hexosaminidase B; D001619:beta-N-Acetylhexosaminidases