title: Effect of 'attenuated' mutations in mucopolysaccharidosis IVA on molecular phenotypes of N-acetylgalactosamine-6-sulfate sulfatase.
abstract: Mucopolysaccharidosis IVA is an autosomal recessive disease caused by the deficiency of N-acetylgalactosamine-6-sulfate sulfatase (GALNS). Mutation screening of the GALNS gene was performed for seven MPS IVA patients with attenuated phenotypes from three unrelated families. Four of 5 missense mutations identified in this study (p.F167V, p.R253W, p.R380S, p.P484S) and two reported (p.F97V, p.N204K), associated with attenuated phenotypes, were characterized using in vitro stable expression experiments, enzyme kinetic study, protein processing and structural analysis. The stably expressed mutant enzymes defining the attenuated phenotype exhibited a considerable residual activity (1.2-36.7% of the wild-type GALNS activity) except for p.R380S. Enzyme kinetic studies showed that p.F97V, p.F167V and p.N204K have lower affinity to the substrate compared with other mutants. The p.F97V enzyme was the most thermolabile at 55 degrees C. Immunoblot analyses indicated a rapid degradation and/or an insufficiency in processing in the mutant proteins. Tertiary structure analysis revealed that although there was a tendency for 'attenuated' mutant residues to be located on the surface of GALNS, they have a different effect on the protein including modification of the hydrophobic core and salt-bridge formation and different potential energy. This study demonstrates that 'attenuated' mutant enzymes are heterogeneous in molecular phenotypes, including biochemical properties and tertiary structure.
authors: Department of Pediatrics, Saint Louis University, Pediatric Research Institute, 3662 Park Ave., St. Louis, MO 63110-2586, USA.
authors: A M
authors: AM
authors: Montaño
authors: K
authors: K
authors: Sukegawa
authors: Z
authors: Z
authors: Kato
authors: R
authors: R
authors: Carrozzo
authors: P
authors: P
authors: Di Natale
authors: E
authors: E
authors: Christensen
authors: K O
authors: KO
authors: Orii
authors: T
authors: T
authors: Orii
authors: N
authors: N
authors: Kondo
authors: S
authors: S
authors: Tomatsu
mesh_terms: D000293:Adolescent; D000328:Adult; D000818:Animals; D016466:CHO Cells; D002811:Chondroitinsulfatases / Q000737:chemistry / Q000172:deficiency / Q000235:genetics* / Q000378:metabolism; D006224:Cricetinae; D003412:Cricetulus; D004252:DNA Mutational Analysis; D004795:Enzyme Stability; D005091:Exons; D005260:Female; D020022:Genetic Predisposition to Disease; D006358:Hot Temperature; D006801:Humans; D057927:Hydrophobic and Hydrophilic Interactions; D007558:Italy; D007564:Japan; D007700:Kinetics; D008297:Male; D008875:Middle Aged; D008958:Models, Molecular; D009085:Mucopolysaccharidosis IV / Q000201:enzymology / Q000235:genetics*; D020125:Mutation, Missense; D010154:Pakistan; D010375:Pedigree; D010641:Phenotype; D011499:Protein Processing, Post-Translational; D017434:Protein Structure, Tertiary; D012720:Severity of Illness Index; D013379:Substrate Specificity; D014162:Transfection
chemical_list: D002811:Chondroitinsulfatases; C102295:GALNS protein, human