title: HFE gene mutations in patients with primary iron overload: is there a significant improvement in molecular diagnosis yield with HFE sequencing?
abstract: Rare HFE variants have been shown to be associated with hereditary hemochromatosis (HH), an iron overload disease. The low frequency of the HFE p.C282Y mutation in HH-affected Brazilian patients may suggest that other HFE-related mutations may also be implicated in the pathogenesis of HH in this population. The main aim was to screen for new HFE mutations in Brazilian individuals with primary iron overload and to investigate their relationship with HH. Fifty Brazilian patients with primary iron overload (transferrin saturation>50% in females and 60% in males) were selected. Subsequent bidirectional sequencing for each HFE exon was performed. The effect of HFE mutations on protein structure were analyzed by molecular dynamics simulation and free binding energy calculations. p.C282Y in homozygosis or in heterozygosis with p.H63D were the most frequent genotypic combinations associated with HH in our sample population (present in 17 individuals, 34%). Thirty-six (72.0%) out of the 50 individuals presented at least one HFE mutation. The most frequent genotype associated with HH was the homozygous p.C282Y mutation (n=11, 22.0%). One novel mutation (p.V256I) was indentified in heterozygosis with the p.H63D mutation. In silico modeling analysis of protein behavior indicated that the p.V256I mutation does not reduce the binding affinity between HFE and β2-microglobulin (β2M) in the same way the p.C282Y mutation does compared with the native HFE protein. In conclusion, screening of HFE through direct sequencing, as compared to p.C282Y/p.H63D genotyping, was not able to increase the molecular diagnosis yield of HH. The novel p.V256I mutation could not be implicated in the molecular basis of the HH phenotype, although its role cannot be completely excluded in HH-phenotype development. Our molecular modeling analysis can help in the analysis of novel, previously undescribed, HFE mutations.
authors.affiliation: Department of Clinical Chemistry and Toxicology, School of Pharmaceutical Sciences, University of Sao Paulo, SP, Brazil. pacaleb@usp.br
authors.forename: Paulo C J L
authors.initials: PC
authors.lastname: Santos
authors.forename: Alexandre C
authors.initials: AC
authors.lastname: Pereira
authors.forename: Rodolfo D
authors.initials: RD
authors.lastname: Cançado
authors.forename: Isolmar T
authors.initials: IT
authors.lastname: Schettert
authors.forename: Tiago J P
authors.initials: TJ
authors.lastname: Sobreira
authors.forename: Paulo S L
authors.initials: PS
authors.lastname: Oliveira
authors.forename: Rosario D C
authors.initials: RD
authors.lastname: Hirata
authors.forename: Mario H
authors.initials: MH
authors.lastname: Hirata
authors.forename: Maria Stella
authors.initials: MS
authors.lastname: Figueiredo
authors.forename: Carlos S
authors.initials: CS
authors.lastname: Chiattone
authors.forename: Jose E
authors.initials: JE
authors.lastname: Krieger
authors.forename: Elvira M
authors.initials: EM
authors.lastname: Guerra-Shinohara
mesh_terms: D001938:Brazil / Q000453:epidemiology; D005820:Genetic Testing; D000071020:Hemochromatosis Protein; D015395:Histocompatibility Antigens Class I / Q000737:chemistry / Q000235:genetics*; D006801:Humans; D019190:Iron Overload / Q000235:genetics*; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics*; D008958:Models, Molecular; D009154:Mutation; D020125:Mutation, Missense; D057089:Pathology, Molecular / Q000379:methods*; D011487:Protein Conformation; D017422:Sequence Analysis, DNA
chemical_list: C103673:HFE protein, human; D000071020:Hemochromatosis Protein; D015395:Histocompatibility Antigens Class I; D008565:Membrane Proteins