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AKU: From classical genetics
to gene cloning

Classical genetics
– Garrod - inborn errors of metabolism (1908)
• alkaptonuria: accumulation of
homogentisic acid in urine
• congenital “inborn” defect
• rare but increased frequency with
consanguinity
–autosomal recessive
AKU: From classical genetics
to gene cloning

Biochemical genetics
– Enzymatic deficiency of AKU demonstrated
in 1958
• Homogentisate 1,2 dioxygenase (HGO)

Mapping
– Trait mapped by homozygosity mapping
and by comparative mapping (mouse gene
on chr 16, sharing conserved synteny with
human 3q)
AKU: From classical genetics
to gene cloning

Functional cloning - getting there
– fungal hmgA was starting point to search
human EST databse
• retrieved sequence and clone
• sequenced clone - encoded peptide with
52% identify with fungal hmgA
• bacterially expressed protein had HGO
activity
• expressed in human liver and kidney
AKU: From classical genetics
to gene cloning

Functional cloning - proving it’s there
– retrieved genomic clone
– HGO mapped by somatic cell hybrid and
FISH analysis - HGO maps to AKU!
– mutations found in affected patients
– mutant protein lacks HGO activity