GLUCOSE PHOSPHATE ISOMERASE DEFICIENCY - BIOCHEMICAL AND MOLECULAR-GENETIC STUDIES ON THE ENZYME VARIANTS OF 2 PATIENTS WITH SEVERE HEMOLYTIC-ANEMIA

Biochemical and molecular genetic studies were performed on the enzyme variants of two patients compound heterozygous for glucose phosphate isomerase (GPI) deficiency, both suffering from severe haemolytic anae mia. The enzymes of case 1 (GPI 'Zwickau') and case 2 (GPI 'Nordhorn' [25]), revealed reduced GPI activity and remarkable thermolability. Gl ucose-6-phosphate (Gluc-6-P) concentration was elevated 2.3 times in c ase 1 and 3.8 times in case 2. Sequencing the patients' GPI genes show ed four different point mutations, two of them involving highly conser ved amino acids. The c1039 C --> T substitution, found in the gene of GPI 'Zwickau', has been described recently [30] and causes an Arg 347 --> Cys substitution close to the putative catalytic site. The second mutation in this case is a novel c1538 G --> A substitution causing a Trp --> stop mutation at position 513 apparently resulting in prematur e RNA degradation thus resulting either in a complete lack of protein or a protein which does not show GPI activity. In the gene of GPI 'Nor dhorn' a c1028 A --> G mutation was discovered, also previously descri bed [1, 9] causing a Gin 343 --> Trp substitution. The second mutation was a novel splice site mutation at the border of intron 15 to exon 1 6: IVS15-(-2) A --> C which leads to an aberrant splicing of exon 16, thus resulting either in a truncated and most likely inactive enzyme o r in no protein at all. Conclusion Biochemical and molecular genetic s tudies performed with the enzyme variants GPI 'Zwickau' and GPI 'Nordh orn' showed that in both cases the simultaneous occurrence of st singl e amino acid substitution affecting the active site, together with a n onsense mutation leading to the loss of major parts of the enzyme prob ably explains the severe clinical course of the disease.