title: A single amino acid change, A91V, leads to conformational changes that can impair processing to the active form of perforin.
abstract: Mutations in the perforin gene have been found in patients with hemophagocytic lymphohistiocytosis (HLH), a rare autosomal recessive disease. We describe a patient expressing perforin with amino acid changes A91V and W374X. The ability of cytotoxic T lymphocytes (CTLs) and natural killer (NK) cells to lyse target cells is greatly reduced. Here we demonstrate that perforin from this patient is not recognized using an antibody raised against native perforin (deltaG9), but is readily detected using an antibody raised against a peptide epitope (2d4), suggesting that the epitope recognized by deltaG9 is destroyed by the change at A91V. Immunoblotting reveals no protein corresponding to the truncated transcript encoded by W374X, revealing that only perforin with the A91V change is expressed in CTLs from the patient. Patient CTLs show bands corresponding to the immature and intermediate forms of perforin, but the mature active form of perforin is absent or barely detectable. The conformational changes and impaired cleavage of A91V perforin are likely to explain the reduced cytotoxicity in CTLs and NK cells from this patient and are likely to contribute to the pathogenesis of HLH.
authors.affiliation: Sir William Dunn School of Pathology, South Parks Rd, Oxford OX1 3RE, United Kingdom.
authors.forename: Christina
authors.initials: C
authors.lastname: Trambas
authors.forename: Federico
authors.initials: F
authors.lastname: Gallo
authors.forename: Daniela
authors.initials: D
authors.lastname: Pende
authors.forename: Stefania
authors.initials: S
authors.lastname: Marcenaro
authors.forename: Lorenzo
authors.initials: L
authors.lastname: Moretta
authors.forename: Carmela
authors.initials: C
authors.lastname: De Fusco
authors.forename: Alessandra
authors.initials: A
authors.lastname: Santoro
authors.forename: Luigi
authors.initials: L
authors.lastname: Notarangelo
authors.forename: Maurizio
authors.initials: M
authors.lastname: Arico
authors.forename: Gillian M
authors.initials: GM
authors.lastname: Griffiths
mesh_terms: D002675:Child, Preschool; D003602:Cytotoxicity, Immunologic / Q000235:genetics; D000939:Epitopes / Q000235:genetics; D005260:Female; D015616:Histiocytosis, Non-Langerhans-Cell / Q000209:etiology / Q000235:genetics; D006801:Humans; D007694:Killer Cells, Natural / Q000276:immunology; D008562:Membrane Glycoproteins / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D020125:Mutation, Missense; D054353:Perforin; D052899:Pore Forming Cytotoxic Proteins; D011487:Protein Conformation; D011499:Protein Processing, Post-Translational; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology
chemical_list: D000939:Epitopes; D008562:Membrane Glycoproteins; D052899:Pore Forming Cytotoxic Proteins; D054353:Perforin