The Structure of Human GALNS Reveals the Molecular Basis for Mucopolysaccharidosis IV A

Abstract

Lysosomal enzymes catalyze the breakdown of macromolecules in the cell. In humans, loss of activity of a lysosomal enzyme leads to an inherited metabolic defect known as a lysosomal storage disorder. The human lysosomal enzyme galactosamine-6-sulfatase (GALNS, also known as N-acetylgalactosamine-6-sulfatase and GalN6S; E.C. 3.1.6.4) is deficient in patients with the lysosomal storage disease mucopolysaccharidosis IV A (also known as MPS IV A and Morquio A). Here we report the three-dimensional structure of human GALNS, determined by x-ray crystallography at 2.2 Å resolution. The structure reveals a catalytic gem diol nucleophile derived from modification of a cysteine side chain. The active site of GALNS is a large, positively charged trench suitable for binding polyanionic substrates such as keratan sulfate and chondroitin-6-sulfate. Enzymatic assays on the insect cell-expressed human GALNS indicate activity against synthetic substrates and inhibition by both substrate and product. Mapping 120 MPS IV A missense mutations onto the structure reveals that a majority of mutations affect the hydrophobic core of the structure, indicating that most MPS IV A cases result from misfolding of GALNS. Comparison of the structure of GALNS to paralogous sulfatases shows a wide variety of active site geometries in the family, but strict conservation of the catalytic machinery. Overall, the structure and the known mutations establish the molecular basis for MPS IV A and for the larger MPS family of diseases.

In silico analysis of mutations occurring in the protein N-acetylgalactosamine-6-sulfatase (GALNS) and causing mucopolysaccharidosis IVA

“…Nacetylgalactosamine-6-sulfatase (GALNS) is the enzyme responsible for the first step in the degradation process of the glycosaminoglycans (GAG) called keratan sulfate (KS) and chondroitin-6-sulfate (C6S). It hydrolyzes the bond between GAG and the sulfate group linked to the 6th carbon of the saccharides, and its deficiency leads to the accumulation of KS and C6S in lysosomes, causing a cellular dysfunction known as mucopolysaccharidosis (MPV) IV, or Morquio syndrome (MS) (Pajares et al, 2012;Rivera-Colón et al, 2012;Kubaski et al, 2013).…”

“…However, the correlation between genotypes and phenotypes is not yet clearly understood (Tomatsu et al, 2011;Rivera-Colón et al, 2012;Pajares et al, 2012;Kubaski et al, 2013).…”

“…It is difficult to estimate the prevalence of MS, due to shortages of basic population studies and epidemiological data, which is estimated to be between 1 in 40,000 and 1 in 50,000 births (Northover et al, 1996;Nelson, 1997;Rivera-Colón et al, 2012). In Brazil, only the state of Rio Grande do Sul has data on the incidence of MS, showing 1 in every 275,000 live births (Schwartz et al, 2008).…”

“…The most often reported mutations of GALNS enzyme that cause MS are classified in three categories: changes in the catalytic site, changes in the hydrophobic core and changes in enzyme surface (Tomatsu et al, 2004;Rivera-Colón et al, 2012). Tomatsu et al (2005) reported a total of 148 mutations, with 103 of them being missense mutations, a number recently revised by Rivera-Colón et al (2012) to 129 missense mutations.…”

“…Tomatsu et al (2005) reported a total of 148 mutations, with 103 of them being missense mutations, a number recently revised by Rivera-Colón et al (2012) to 129 missense mutations. Since the tertiary structure of the GALNS enzyme is known (Rivera-Colón et al, 2012), the search for possible regions of intrinsic disorder in the mutant protein has substantial importance to enable protein modeling and virtual screening assays of these mutants, allowing the discovery of molecules with theoretic binding potential to the enzyme, and eventual pharmacological reactivation of the affected enzyme.…”