title: Novel mutations and structural implications in R-type pyruvate kinase-deficient patients from Southern Italy.
abstract: Deficiency of the R-type pyruvate kinase (R-PK) causes an autosomal recessive, hereditary, nonspherocytic hemolytic anemia (HNSHA). We screened seven unrelated patients from the south of Italy for the known mutations and found one patient homozygous for the 1529A (R510Q) mutation, two others bearing the 1456T (R486W) mutation, one homozygous and another heterozygous, and two heterozygotes for the 994A mutation (G332S). We also found three novel mutations at the heterozygote status: a G to C transversion in position 1010 (1010C; R337P) and a C to T transition in position 1492 (1492T; R498C), which are missense, and a T to G transversion in position 1523 (1523G; L508Z), which produces a stop codon with a subsequent loss of the C-terminal protein domain. The structural features of R-PK in the mutation-bearing regions were examined. In all cases the mutations altered the local conformation of the enzyme. Both G332S and R337P are in highly conserved sequence regions. In particular, the R337P mutation significantly affects the intersubunit interactions, because it is located in a region subjected to a large conformational change that occurs during the R-->T allosteric transition, which is essential for the enzyme activity. The R486W mutation affects an external pocketlike region, producing only a local conformational change; the R498C mutation changes the interactions among neighbouring residues; the R510Q mutation involves the loss of interdomain interactions that may reduce enzyme stability and activity. Our data also indicate that in patients from Southern Italy, pyruvate kinase deficiency is heterogeneous, the 1529A mutation, which is the most frequent mutation in the U.S. Caucasian population, having a lower frequency.
authors.affiliation: Dipartimento di Biochimica e Biotecnologie Mediche and CEINGE-Biotecnologie Avanzate, Federico II University, Naples, Italy.
authors.forename: L
authors.initials: L
authors.lastname: Pastore
authors.forename: R
authors.initials: R
authors.lastname: Della Morte
authors.forename: G
authors.initials: G
authors.lastname: Frisso
authors.forename: F
authors.initials: F
authors.lastname: Alfinito
authors.forename: D
authors.initials: D
authors.lastname: Vitale
authors.forename: R M
authors.initials: RM
authors.lastname: Calise
authors.forename: F
authors.initials: F
authors.lastname: Ferraro
authors.forename: A
authors.initials: A
authors.lastname: Zagari
authors.forename: B
authors.initials: B
authors.lastname: Rotoli
authors.forename: F
authors.initials: F
authors.lastname: Salvatore
mesh_terms: D000494:Allosteric Regulation / Q000235:genetics; D000595:Amino Acid Sequence; D000745:Anemia, Hemolytic, Congenital / Q000145:classification / Q000235:genetics*; D017124:Conserved Sequence / Q000235:genetics; D004252:DNA Mutational Analysis; D004795:Enzyme Stability; D006801:Humans; D007558:Italy; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011487:Protein Conformation; D011770:Pyruvate Kinase / Q000172:deficiency* / Q000235:genetics; D012156:Reticulocytes / Q000201:enzymology; D017386:Sequence Homology, Amino Acid
chemical_list: D011770:Pyruvate Kinase