Aims 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, the HMBS gene 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 of HMBS gene mutations in patients with AIP in South Africa. The biochemical characterisation of expressed HMBS mutants reveals insight into the mechanism of catalytic activity loss, which may inspire investigation into individualised therapy based on the molecular lesion identified.
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Handling editor Tahir Pillay
Contributors PM: conceived and oversaw the project. PF and AC: collection and interpretation of patient mutation data. EP and AC: enzyme expression; collection and interpretation of enzyme kinetic data. MS and CWS: clinical management of AIP patients at Groote Schuur Hospital. The first draft of the manuscript was written by PF and critically reviewed by all authors.
Funding Project funding was provided by the Medical Research Council (self-initiated research grant awarded to PM), the National Research Foundation Incentive Award for rated researchers (PM) and the University of Cape Town Research Committee.
Competing interests None.
Ethics approval Health Sciences Faculty Research Ethics Committee (HREC/REF: 324/2010), University of Cape Town, South Africa.
Provenance and peer review Not commissioned; externally peer reviewed.
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