Pyruvate kinase “Göttingen_1,2”: Congenital hemolytic anemia, evidence of double heterozygosity, and lack of enzyme cooperativity

Summary

Double heterozygosity of pyruvate kinase (PK) deficiency associated with hereditary hemolytic anemia is emphasized by studies of a kindred harboring two distinct mutant forms of this enzyme. The hematologically unaffected parents exhibit slightly reduced PK activity, a normal Hill coefficient, and a normal thermodynamic dissociation constant for the overall reaction. The paternal enzyme is characterized by normal substrate affinities and decreased activities with the substrate analogues CDP and GDP, whereas the maternal enzyme shows normal affinity for PEP, but an increased affinity for ADP and low thermostability. It is assumed that the erythrocytes of the parents contain a mixture of normal PK and a functionally abnormal isoenzyme, the latter differing between the parents. The two children suffer from hereditary hemolytic anemia. Their PK must be a combination of the mutant paternal and maternal isoenzymes, and their activities are reduced to about 30%. These enzymes are characterized by an increased affinity for PEP and a decreased affinity for ADP, a Hill coefficient of about 1 (indicating lack of cooperativity due to a loss of its allosteric properties), a decreased overall catalytic activity, and a higher resistance to heat denaturation. Further differences are observed in the SDS-gel electrophoresis between the two patients' enzymes. From the enzymological point of view it is impossible to characterize true PK variants in such double heterozygous cases which contain a combination of two different isoenzymes. The cause of chronic hemolysis appears to depend mainly on the loss of the allosteric properties, i.e., the lack of enzyme cooperativity.