title: Molecular characterisation of acute intermittent porphyria in a cohort of South African patients and kinetic analysis of two expressed mutants.
abstract: OBJECTIVE
  Acute intermittent porphyria (AIP) is a disorder of the haem biosynthetic pathway caused by mutations in the hydroxymethylbilane synthase (HMBS) gene. Knowledge of the spectrum of mutations present in South Africa is limited. This study presents the molecular profile of 20 South African patients with AIP, and the kinetic analysis of one novel expressed mutated HMBS enzyme and a previously identified mutation at the same position.
  
  
  METHODS
  Genomic DNA was isolated from affected probands and selected family members, theHMBSgene amplified and mutations characterised by direct sequencing and restriction enzyme analysis. One of the novel mutations (p.Lys98Glu), a previously characterised mutation at the same position (p.Lys98Arg), and the wild-type enzyme were expressed, purified and subjected to partial kinetic characterisation.
  
  
  RESULTS
  Four new mutations, p.Lys98Glu, p.Asp230Aspfs*20, c.161-1G>A and c.422+3_6delAAGT, are described. Seven previously described mutations were found, while four patients revealed no mutations. Mutation analysis of five offspring of one of the probands carrying the p.Trp283X mutation revealed two asymptomatic carriers. Kinetic analysis showed that the p.Lys98Glu mutation results in loss of substrate affinity, whereas the previously described p.Lys98Arg mutation causes the loss of binding between the enzyme and its dipyrromethane cofactor, rendering the enzyme inactive.
  
  
  CONCLUSIONS
  This study comprises the most comprehensive characterisation ofHMBSgene mutations in patients with AIP in South Africa. The biochemical characterisation of expressedHMBSmutants reveals insight into the mechanism of catalytic activity loss, which may inspire investigation into individualised therapy based on the molecular lesion identified.
authors.affiliation: Division of Chemical Pathology, Department of Pathology, Groote Schuur Hospital, University of Cape Town, Observatory, South Africa.
authors.forename: Philip
authors.initials: P
authors.lastname: Fortgens
authors.affiliation: Division of Medical Biochemistry, Department of Integrative Biomedical Sciences, Lennox Eales Porphyria Laboratory, University of Cape Town, Observatory, South Africa.
authors.forename: Elaine
authors.initials: E
authors.lastname: Pienaar
authors.affiliation: Lennox Eales Porphyria Laboratory, University of Cape Town, Observatory, South Africa.
authors.forename: Anne
authors.initials: A
authors.lastname: Corrigall
authors.affiliation: Division of Hepatology, Department of Medicine, Groote Schuur Hospital, University of Cape Town, Observatory, South Africa.
authors.forename: Mark
authors.initials: M
authors.lastname: Sonderup
authors.affiliation: Division of Hepatology, Department of Medicine, Groote Schuur Hospital, University of Cape Town, Observatory, South Africa.
authors.forename: C Wendy
authors.initials: CW
authors.lastname: Spearman
authors.affiliation: Department of Integrative Biomedical Sciences, Lennox Eales Porphyria Laboratory, Structural Biology Research Unit, University of Cape Town, Observatory, South Africa.
authors.forename: Peter
authors.initials: P
authors.lastname: Meissner
mesh_terms: D044383:Black People / Q000208:ethnology / Q000235:genetics; D015331:Cohort Studies; D005260:Female; D006801:Humans; D011163:Hydroxymethylbilane Synthase / Q000235:genetics*; D008297:Male; D009154:Mutation / Q000235:genetics*; D017118:Porphyria, Acute Intermittent / Q000208:ethnology / Q000235:genetics*; D013019:South Africa / Q000208:ethnology
chemical_list: D011163:Hydroxymethylbilane Synthase
keywords: CHEMICAL PATHOLOGY; ENZYMES; MOLECULAR BIOLOGY