Novel homozygous c.2039C>G (Thr680Ser) missense mutation in HK1 is associated with decreased HK activity.

(A) DNA sequence variations in HK1 in both the patient and a healthy control revealed 5 base changes that represent either polymorphic substitutions or sequence discrepancies and, in addition, 2 postulated polymorphisms (c.78G>C and IVS15+27C>T). A homozygous c.2039C>G mutation is detected only in the patient, and the consequent Thr680Ser substitution affects a highly conserved and structurally important residue in the active site of HK-I. (B) In the family pedigree chart, individuals heterozygous or homozygous for the c.2039C>G mutation are indicated by half-filled and filled symbols, respectively. Lanes in the agarose gel for each family member are directly below the symbol for that individual. The agarose gel below shows a 301-bp PCR product encompassing exon 15, which was amplified from genomic DNA and subjected to AciI digestion. c.2039C>G creates an additional restriction site upon the 2 normally present in this fragment. Thus, digestion of the wild-type allele results in fragments of 14, 221, and 66 bp, whereas the extra AciI recognition sequence yields additional fragments of 34 and 187 bp. Digestion fragments are indicated by arrows on the right (14 and 34 bp, fragments not shown). Homozygosity for c.2039C>G is confirmed in patient Z62. The patient's father (Z32), mother (H37), sisters (Z53, Z56, and Z66.)15and daughter N84 were all heterozygous, whereas sister Z56 didn't carry the mutation. All family members heterozygous for the Thr680Ser substitution displayed reduced HK activity in their red blood cells, ranging from 0.64-0.89 U/gHb (reference value: 1.34 ± 0.42 U/gHb).15 HK activity for sister Z56. was 1.22 U/gHb. *indicates Z62; M, marker; NC, healthy control.