Molecular Analysis of 29 Pyruvate Kinase–Deficient Patients From Central Europe With Hereditary Hemolytic Anemia

Abstract

We investigated the DNA of 29 unrelated pyruvate kinase (PK) deficiency (PKD) patients from Central Europe with hereditary nonspherocytic hemolytic anemia for mutations in the PK-L/R gene. Among 58 potentially affected alleles, 53 mutations were identified, of which 17 were different from each other. Of these 17 mutations, 13 were single-nucleotide (nt) substitutions resulting in amino acid exchanges, G787A (Gly263-Arg), G994A (Gly332-Ser), G1006T (Ala336-Ser), G1010A (Arg337-Gln), A1081G (Asn361-Asp), G1127T (Ser376-Ile), G1174A (Ala392-Thr), G1281T (Glu427-Asp), C1454T (Ser485-Phe), C1456T (Arg486-Trp), G1493A (Arg498-His), G1529A (Arg510-Gln), and C1594T (Arg532-Trp); 1 in-frame triplet deletion, 1060delAAG (delLys354); 1 in-frame triplet insertion, 1203insAGC (insSer after Cys401); 1 splicesite mutation, 101-1G-A; and 1 frameshift deletion, 628delGT. Six mutations, 628delGT, G787A, G1010A, G1127T, G1281T, and C1454T, are described for the first time. To test the hypothesis of a single origin of the most common PK mutation in the European population, G1529A, we investigated all patients at four polymorphic sites in the PK-L/R gene: C/A at nt 1705, C/T at nt 1992, the (ATT)n microsatellite in intron J, and a polymorphism (T)10/(T)19 in intron I. Nine patients homozygous for mutation G1529A were consistent in all four markers. In the group of patients homozygous for mutation G1529A, the hematologic parameters and clinical manifestations have been studied in detail. Although having an identical mutation in the PK-L/R gene, the patients are affected differently. Their appearance ranges from a very mild compensated hemolysis to a severe anemia. Possible molecular explanations are discussed.

Molecular characterization of the PK‐LR gene in pyruvate kinase deficient Spanish patients

“…In patient 1 the nonsense mutation Arg559 creates a premature stop and therefore a truncated protein with abnormal enzyme activity. In patients 2 and 7 the missense mutation Gln337 (1010A), previously described by Lenzner et al (1997 ), is located near the inorganic cofactors Mn +2 ligands which may be altered. This specific mutation is characterized by severe clinical manifestations as demonstrated by the existence of haemolysis in the heterozygous condition (patient 7).…”

“…The cloning of the R‐type cDNA has enabled the study of this enzymopathy at the DNA level ( Tani et al , 1988 ; Kanno et al , 1991 ), and, up to now, about 80 mutations involved in the deficient PK‐LR gene have been described ( Baronciani et al , 1996 ; Baronciani & Beutler, 1993a, b; Kanno et al , 1991 , 1992, 1993a, b, c; Lakomek et al , 1994 ; Lenzner et al , 1994 , 1997; Neubauer et al , 1991 ; Bianchi et al , 1994 , 1995; Rouger et al , 1996 ). This is the first report of a complete mutation analysis of the PK‐LR gene performed in 12 unrelated PKD patients from Spain.…”

Residual pyruvate kinase activity in PKLR‐deficient erythroid precursors of a patient suffering from severe haemolytic anaemia

“…17 In several cases, however, persistence of PKM2 in RBC of PK-R-deficient patients is reported. 8,9,18 We, therefore, assessed PKM2 expression in patient-derived reticulocytes and cultured erythroblasts. We found these progenitors to express levels of PKM2 comparable with control reticulocytes and erythroblasts ( Figure 3B).…”

Red cell pyruvate kinase deficiency: molecular and clinical aspects

“…Three of the most prevalent mutations in patients with PK deficiency are 1529A, 1456T and 1468T; these variants have been found to be distributed with a strong ethnic and regional background. In particular, the 1529A is the most common mutation in the USA (42%) (Baronciani & Beutler, 1995) and in Northern and Central Europe (41%) (Lenzner et al, 1997a).…”