Crystal Structure of β-Hexosaminidase B in Complex with Pyrimethamine, a Potential Pharmacological Chaperone

Abstract

Abstractβ-Hexosaminidases (β-hex) are a group of glycosyl hydrolase isozymes that break down neutral and sialylated glycosphingolipids in the lysosomes, thereby preventing their buildup in neuronal cells. Some mutants of β-hex have decreased folding stability that results in adult-onset forms of lysosomal storage diseases. However, prevention of the harmful accumulation of glycolipids only requires 10% of wild-type activity. Pyrimethamine (PYR) is a potential pharmacological chaperone that works by stabilizing these mutant enzymes sufficiently to allow more β-hex to arrive in the lysosome, where it can carry out its function. An X-ray structure of the complex between human β-hexosaminidase B (HexB) and PYR has been determined to 2.8 Å. PYR binds to the active site of HexB where several favorable van der Waals contacts and hydrogen bonds are † Accession Codes PDB code: 3LMY.

Pyrimethamine Derivatives: Insight into Binding Mechanism and Improved Enhancement of Mutant β-N-acetylhexosaminidase Activity

“…Removal of one (cf. 8,9) or both (cf. 11) amino groups from the pyrimidine ring resulted in a >10-fold increase in IC 50 values ( Table 1).…”

“…This is consistent with their role as H-bond donors within the active site of the enzyme deduced from the 3D structure of the HexB:PYR complex. 9 Inhibitory Activity of PYR Analogues with Modified 6-Alkyl Side Chains. Shortening of the alkyl side chain from an ethyl to methyl side arm resulted in a more than 25-fold increase in IC 50 values, i.e., from 17 to 560 μM (Table 2).…”

“…2 The structure of Hex B in complex with PYR has only recently been elucidated. 9 Amino acids Asp240 and Glu491 are within hydrogen-bonding distances of the amino groups in PYR and are predicted to act as hydrogen bond acceptors ( Figure 1B). The ethyl group of PYR is positioned within a hydrophobic pocket formed by Trp405, Trp424, and Trp489.…”

From structural biology to designing therapy for inborn errors of metabolism

“…To date, PCs have promising potentials for several IEMs, including the Fabry, Gaucher and Pompe Diseases, which have reached early-stage clinical trials (Boyd et al 2013; Parenti et al 2015); while others are gaining proof of concept (Santos-Sierra et al 2012; Jorge-Finnigan et al 2013; Makley et al 2015). These first-generation PC molecules often originate as substrate-mimetics or cofactor-mimetics of the target enzymes, and the field of structural biology has been useful in characterizing how the PC molecule interacts with the enzyme active site (Bateman et al 2011; Guce et al 2011; Torreblanca et al 2012; Suzuki et al 2014). However, active-site-directed PCs create a conundrum in which by binding to the active site, these ligands could potentially compete with the native substrate or cofactor of the target enzyme.…”

Evidence that small molecule enhancement of β-hexosaminidase activity corrects the behavioral phenotype in Dutch APPE693Q mice through reduction of ganglioside-bound Aβ

“…35 Pyrimethamine, like the iminosugar PC NAG-thiazoline, 36 was shown to bind the active site of β-hex and rescue certain β-hex A mutants that cause Tay–Sachs disease or Sandhoff disease. 37 In a phase 1/2 trial in Tay–Sachs and Sandhoff patients, pyrimethamine treatment increased β-hex A levels for all eight patients; however, severe side effects prevented further development. 33 Carbohydrate mimetics including iminosugars like NAG-thiazoline are more potent and would likely demonstrate better tolerability.…”