Chapter 12, Problem 36P

Alkaptonuria is a human autosomal recessive disorder caused by mutation of the HAO gene that encodes the enzyme homogentisic acid oxidase. Restriction mapping

of the HAO gene region reveals four BamHI restriction sites $\left(\text{B1 to B4}\right)$ in the wild $\text{-}$ type allele and three BamHI restriction sites in the mutant allele. BamHI utilizes the restriction sequence $\text{5'-GGATCC-3'}$. The BamHI restriction sequence identified as $\text{B3}$ is altered to $\text{5'-GGAACC-3'}$ in the mutant allele. The mutation results in a $\text{Ser-to-Thr}$ missense mutation. Restriction maps of the two alleles are shown below, and the binding sites of two molecular probes (probe A and probe B) are identified.

DNA samples taken from a mother (M), father (F), and two children $\left(\text{C1 and C2}\right)$ are analyzed by Southern blotting of $\text{BamHI-digested DNA}$. The gel electrophoresis results are illustrated.

a. Using A to represent the $\text{wild-type}$ allele and a for the mutant allele, identify the genotype of each family member. Identify any family member who is alkaptonuric.

b. In a separate figure, draw the autoradiograph patterns for all the genotypes that could be found in children of this couple.

c. Explain how the DNA sequence change results in a $\text{Ser-to-Thr}$ missense mutation.