title: Molecular and functional characterization of a new X-linked chronic granulomatous disease variant (X91+) case with a double missense mutation in the cytosolic gp91phox C-terminal tail.
abstract: We report here two atypical cases of X-linked CGD patients (first cousins) in which cytochrome b(558) is present at a normal level but is not functional (X91+). The mutations were localized by single-strand conformational polymorphism of reverse transcriptase-polymerase chain reaction amplified fragments and then identified by sequence analysis. They consisted in two base substitutions (C919 to A and C923 to G), changing His303 to Asn and Pro304 to Arg in the cytosolic gp91phox C-terminal tail. Mismatched polymerase chain reaction and genomic DNA sequencing showed that mothers had both wild-type and mutated alleles, confirming that this case was transmitted in an X-linked fashion. A normal amount of FAD was found in neutrophil membranes, both in the X91+ patients and their parents. Epstein-Barr virus-transformed B lymphocytes from the X91+ patients acidified normally upon stimulation with arachidonic acid, indicating that the mutated gp91phox still functioned as a proton channel. A cell-free translocation assay demonstrated that the association of the cytosolic factors p47phox and p67phox with the membrane fraction was strongly disrupted. We concluded that residues 303 and 304 are crucial for the stable assembly of the NADPH oxidase complex and for electron transfer, but not for its proton channel activity.
authors: GREPI EA 2938 UJF, Laboratoire d'Enzymologie, CHU 38043 Grenoble Cedex 9, France. mjstasia@chu-grenoble.fr
authors: Marie José
authors: MJ
authors: Stasia
authors: Bernard
authors: B
authors: Lardy
authors: Andres
authors: A
authors: Maturana
authors: Pascale
authors: P
authors: Rousseau
authors: Cécile
authors: C
authors: Martel
authors: Pierre
authors: P
authors: Bordigoni
authors: Nicolas
authors: N
authors: Demaurex
authors: Françoise
authors: F
authors: Morel
mesh_terms: D002462:Cell Membrane / Q000378:metabolism; D003573:Cytochrome b Group / Q000378:metabolism; D003600:Cytosol / Q000378:metabolism; D005182:Flavin-Adenine Dinucleotide / Q000032:analysis; D006105:Granulomatous Disease, Chronic / Q000097:blood / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D007223:Infant; D008297:Male; D008562:Membrane Glycoproteins / Q000235:genetics* / Q000378:metabolism; D020125:Mutation, Missense; D009240:N-Formylmethionine Leucyl-Phenylalanine; D000074662:NADPH Oxidase 2; D019255:NADPH Oxidases / Q000378:metabolism; D009504:Neutrophils / Q000201:enzymology; D018807:Polymorphism, Single-Stranded Conformational; D012333:RNA, Messenger / Q000378:metabolism; D013755:Tetradecanoylphorbol Acetate
chemical_list: D003573:Cytochrome b Group; D008562:Membrane Glycoproteins; D012333:RNA, Messenger; D005182:Flavin-Adenine Dinucleotide; D009240:N-Formylmethionine Leucyl-Phenylalanine; C036888:cytochrome b558; C056558:CYBB protein, human; D000074662:NADPH Oxidase 2; D019255:NADPH Oxidases; D013755:Tetradecanoylphorbol Acetate