title: Molecular and functional analysis of the C-terminal region of human erythroid-specific 5-aminolevulinic synthase associated with X-linked dominant protoporphyria (XLDPP).
abstract: Frameshift mutations in the last coding exon of the 5-aminolevulinate synthase (ALAS) 2 gene were described to activate the enzyme causing increased levels of zinc- and metal-free protoporphyrin in patients with X-linked dominant protoporphyria (XLDPP). Only two such so-called gain-of-function mutations have been reported since the description of XLDPP in 2008. In this study of four newly identified XLDPP families, we identified two novel ALAS2 gene mutations, a nonsense p.Q548X and a frameshift c.1651-1677del26bp, along with a known mutation (delAGTG) found in two unrelated families. Of relevance, a de novo somatic and germinal mosaicism was present in a delAGTG family. Such a phenomenon may explain the high proportion of this mutation in XLDPP worldwide. Enhancements of over 3- and 14-fold in the catalytic rate and specificity constant of purified recombinant XLDPP variants in relation to those of wild-type ALAS2 confirmed the gain of function ascribed to these enzymes. The fact that both p.Q548X and c.1651-1677del26bp are located in close proximity and upstream from the two previously described mutations led us to propose the presence of a large gain-of-function domain within the C-terminus of ALAS2. To test this hypothesis, we generated four additional nonsense mutants (p.A539X, p.G544X, p.G576X and p.V583X) surrounding the human XLDPP mutations and defined an ALAS2 gain-of-function domain with a minimal size of 33 amino acids. The identification of this gain-of-function domain provides important information on the enzymatic activity of ALAS2, which was proposed to be constitutively inhibited, either directly or indirectly, through its own C-terminus.
authors.affiliation: AP-HP, Centre Franc¸ais des Porphyries, Hoˆ pital Louis Mourier, 178 rue des Renouillers, 92701 Colombes Cedex,France.
authors.forename: Sarah
authors.initials: S
authors.lastname: Ducamp
authors.forename: Xiaoye
authors.initials: X
authors.lastname: Schneider-Yin
authors.forename: Felix
authors.initials: F
authors.lastname: de Rooij
authors.forename: Jerome
authors.initials: J
authors.lastname: Clayton
authors.forename: Erica J
authors.initials: EJ
authors.lastname: Fratz
authors.forename: Alice
authors.initials: A
authors.lastname: Rudd
authors.forename: George
authors.initials: G
authors.lastname: Ostapowicz
authors.forename: George
authors.initials: G
authors.lastname: Varigos
authors.forename: Thibaud
authors.initials: T
authors.lastname: Lefebvre
authors.forename: Jean-Charles
authors.initials: JC
authors.lastname: Deybach
authors.forename: Laurent
authors.initials: L
authors.lastname: Gouya
authors.forename: Paul
authors.initials: P
authors.lastname: Wilson
authors.forename: Gloria C
authors.initials: GC
authors.lastname: Ferreira
authors.forename: Elisabeth I
authors.initials: EI
authors.lastname: Minder
authors.forename: Hervé
authors.initials: H
authors.lastname: Puy
mesh_terms: D000624:5-Aminolevulinate Synthetase / Q000737:chemistry / Q000172:deficiency / Q000235:genetics* / Q000378:metabolism; D000595:Amino Acid Sequence; D001483:Base Sequence; D002675:Child, Preschool; D018389:Codon, Nonsense; D004252:DNA Mutational Analysis; D005091:Exons; D005260:Female; D016368:Frameshift Mutation; D056726:Genetic Association Studies; D040181:Genetic Diseases, X-Linked / Q000201:enzymology / Q000235:genetics*; D006801:Humans; D007223:Infant; D007700:Kinetics; D008969:Molecular Sequence Data; D009030:Mosaicism; D016297:Mutagenesis, Site-Directed; D010375:Pedigree; D017434:Protein Structure, Tertiary; D046351:Protoporphyria, Erythropoietic / Q000201:enzymology / Q000235:genetics*; D017422:Sequence Analysis, DNA; D055815:Young Adult
chemical_list: D018389:Codon, Nonsense; D000624:5-Aminolevulinate Synthetase; C480079:ALAS2 protein, human