title: Compound heterozygous mutations affect protein folding and function in patients with congenital sucrase-isomaltase deficiency.
abstract: OBJECTIVE
  Congenital sucrase-isomaltase (SI) deficiency is an autosomal-recessive intestinal disorder characterized by a drastic reduction or absence of sucrase and isomaltase activities. Previous studies have indicated that single mutations underlie individual phenotypes of the disease. We investigated whether compound heterozygous mutations, observed in some patients, have a role in disease pathogenesis.
  
  
  METHODS
  We introduced mutations into the SI complementary DNA that resulted in the amino acid substitutions V577G and G1073D (heterozygous mutations found in one group of patients) or C1229Y and F1745C (heterozygous mutations found in another group). The mutant genes were expressed transiently, alone or in combination, in COS cells and the effects were assessed at the protein, structural, and subcellular levels.
  
  
  RESULTS
  The mutants SI-V577G, SI-G1073D, and SI-F1745C were misfolded and could not exit the endoplasmic reticulum, whereas SI-C1229Y was transported only to the Golgi apparatus. Co-expression of mutants found on each SI allele in patients did not alter the protein's biosynthetic features or improve its enzymatic activity. Importantly, the mutations C1229Y and F1745C, which lie in the sucrase domains of SI, prevented its targeting to the cell's apical membrane but did not affect protein folding or isomaltase activity.
  
  
  CONCLUSIONS
  Compound heterozygosity is a novel pathogenic mechanism of congenital SI deficiency. The effects of mutations in the sucrase domain of SIC1229Y and SIF1745C indicate the importance of a direct interaction between isomaltase and sucrose and the role of sucrose as an intermolecular chaperone in the intracellular transport of SI.
authors.affiliation: Department of Physiological Chemistry, University of Veterinary Medicine, Hannover, Germany.
authors.forename: Marwan
authors.initials: M
authors.lastname: Alfalah
authors.forename: Markus
authors.initials: M
authors.lastname: Keiser
authors.forename: Tosso
authors.initials: T
authors.lastname: Leeb
authors.forename: Klaus-Peter
authors.initials: KP
authors.lastname: Zimmer
authors.forename: Hassan Y
authors.initials: HY
authors.lastname: Naim
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D019556:COS Cells; D002239:Carbohydrate Metabolism, Inborn Errors / Q000235:genetics* / Q000378:metabolism*; D002522:Chlorocebus aethiops; D004789:Enzyme Activation / Q000235:genetics; D015971:Gene Expression Regulation, Enzymologic; D018740:Genetic Heterogeneity; D006801:Humans; D016297:Mutagenesis, Site-Directed; D017354:Point Mutation; D017510:Protein Folding; D017434:Protein Structure, Tertiary; D021381:Protein Transport / Q000235:genetics; D013394:Sucrase-Isomaltase Complex / Q000737:chemistry / Q000172:deficiency / Q000235:genetics; D013395:Sucrose / Q000378:metabolism*; D014162:Transfection
chemical_list: D013395:Sucrose; D013394:Sucrase-Isomaltase Complex