Seven Pyruvate Kinase Variants Characterized by the ICSH Recommended Methods

Shiro, Motoo; Fujii, H.; Takegawa, Susumu; Nakatsuji, T.; Yamato, Kunio; Ishida, Yoshiteru; Ninomiya, Nobuyuki

doi:10.1111/j.1365-2141.1980.tb07181.x

Cited by 50 publications

(23 citation statements)

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“…We have previously reported the clinical history and laboratory data of patients YK and FK (13). This variant enzyme was characterized by increased K0.5S (phosphoenolpyruvate), thermal instability, and slow electrophoretic mobility (13). Because the slow electrophoretic mobility of this variant PK had been demonstrated in both erythrocytes and liver (14), a structural mutation within the region common to both L-and R-type PKs was expected.…”

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confidence: 99%

“…Here, we report the complete nucleotide sequence and the deduced amino acid sequence of human R-type PK ¶ and also present evidence of a PK variant (PK Tokyo) caused by a single amino acid substitution in R-type PK. We have previously reported the clinical history and laboratory data of patients YK and FK (13). This variant enzyme was characterized by increased K0.5S (phosphoenolpyruvate), thermal instability, and slow electrophoretic mobility (13).…”

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confidence: 99%

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cDNA cloning of human R-type pyruvate kinase and identification of a single amino acid substitution (Thr384----Met) affecting enzymatic stability in a pyruvate kinase variant (PK Tokyo) associated with hereditary hemolytic anemia.

Kanno¹,

Fujii²,

Hirono³

et al. 1991

Proc. Natl. Acad. Sci. U.S.A.

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ABSTRACTcDNA clones for human R-type pyruvate kinase (PK) were isolated from a human reticulocyte cDNA library, constructed by PCR with a single gene-spedflc primer. The full-length cDNA was 2060 base pairs long, and the cDNA encoded 574 amino adds, the same number as that by rat R-type PK. Compared with human L-type PK, R-type PK was 31 amino acids longer at the amino terminus. We also cloned and characterized R-type PK cDNA clones from patients with hereditary hemolytic anemia from a PK deficiency, PK Tokyo. A single nucleotide substitution (ACG to ATG) was found at nucleotide 1151 of the coding sequence of the R-type PK, which caused an amino acid substitution, Thr3N -Met. Dot blot hybridization of PCR-ampifled genomic DNA from patients and their parents by allele-specific oligonucleotide probes showed that the patients were homozygous for this nucleotide change and that the parents, who were second cousins, were heterozygous. To confirm that the nucleotide change was responsible for the variant phenotype, we expressed the L-type PK with the single amino acid change in Escherichia coil and characterized the enzyme. The variant PK was thermolabile and moved slowly in the polyacrylamide gel buffered in 10 mM Tris-HCI, pH 8.3; these characteristics were fully compatible with data obtained from the patient's PK. From these results, we concluded that enzymatic stability of the variant was affected by the point mutation of the PK-encoding gene.

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cDNA cloning of human R-type pyruvate kinase and identification of a single amino acid substitution (Thr384----Met) affecting enzymatic stability in a pyruvate kinase variant (PK Tokyo) associated with hereditary hemolytic anemia.

Kanno¹,

Fujii²,

Hirono³

et al. 1991

Proc. Natl. Acad. Sci. U.S.A.

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“…In almost all cases, family studies confirmed that variant alleles which appeared in a child were also present in the parents. One possible exception to this finding, however, is suggested by family studies of the pyruvate kinase (PK) deficiency in Japanese (Satoh et al 1983;Miwa et al 1980;Ishida et al 1981). PK deficiency causes an anemia, which is associated with null activity in Caucasians in homozygotes, and no disease, but intermediate enzyme levels in heterozygotes (Valentine, Tanaka and Miwa 1961).…”

Section: Activity Variants and Null Allelesmentioning

confidence: 99%

“…PK deficiency causes an anemia, which is associated with null activity in Caucasians in homozygotes, and no disease, but intermediate enzyme levels in heterozygotes (Valentine, Tanaka and Miwa 1961). In Japanese, at least seven different PK alleles with decreased activity have been identified (Miwa et al 1980). These alleles retain some PK activity, in contrast to the null allele found in Caucasians, and consequently family studies often do not indicate significant reduction of PK activity in either parent of an affected individual.…”

Section: Activity Variants and Null Allelesmentioning

confidence: 99%

The estimate of protein polymorphism in human populations: Lack of evidence for overestimation due to post-translational modification.

Fuerst

Ferrell

1985

The Japanese journal of genetics

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New sensitive population screening procedures, such as isoelectric focusing, enzyme activity assays and the use of multiple electrophoretic conditions have uncovered considerable hidden variation at many structural loci. It has been suggested, however, that a significant portion of the newly identified variation is due to alleles at modifier genes which cause the post-translational modification of the structural gene product. Some investigators have questioned whether genetic variability assigned to structural loci in Drosophila has been greatly overestimated because of a failure to recognize the existence of such polymorphic modifier genes. Data from human populations has been reviewed for evidence of such genetically mediated posttranslational modification (GMPTM).Tnree genetic systems (glucose-6-phosphate dehydrogenase, alpha-1-antitrypsin and hemoglobin) are reviewed in detail. Each system is subject to non-genetic post-translational modification, but none of the data currently available indicates any role of GMPTM.Other data, including studies on null alleles, activity variants, population surveys and somatic cell hybrids is also reviewed. Again, no convincing evidence has been uncovered for the presence of GMPTM. It is concluded that the current estimates of polymorphism for enzymes and structural proteins in human populations have not been inflated due to the presence of polymorphism for modifier loci.

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“…In particular, it has become apparent that pyruvate kinase (PK) deficiency often shows either normal or increased red cell PK activity (Miwa et al 1980). The accumulation of red cell 2,3-diphosphoglycerate (2,3-DPG) and the decrease of adenosine-5'-triphosphate (ATP) are characteristically seen in most cases of PK deficiency.…”

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Recommended methods for an additional red cell enzyme (pyrimidine 5′-nucleotidase) assay and the determination of red cell adenosine-5′-triphosphate, 2,3-diphosphoglycerate and reduced glutathione

Shiro¹,

Luzzatto²,

Rosa³

et al. 1989

Clinical &Amp; Laboratory Haematology

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In 1977 the Red Cell Enzyme Expert Panel of the International Committee for Standardization in Haematology (ICSH) issued recommended methods for red cell enzyme analysis . These methods have made it possible to measure red cell enzyme activity with reliable and convenient techniques, and to compare results obtained in different laboratories. After the publication of the ICSH recommended methods, pyrimidine 5'-nucleotidase (P5N) deficiency came to our attention. This enzyme was first described by Valentine et al. (1974) as a cytosolic enzyme in human red cells. Hereditary deficiency results in nonspherocytic haemolytic anaemia. This syndrome is characterized by haemolytic anaemia, pronounced basophilic stippling of red cells, and marked increase in both red cell reduced glutathione (GSH) and pyrimidine-containing nucleotides. To date, more than 40 cases have been reported (Miwa & Fujii 1985). P5N deficiency appears to be one of the more common causes of hereditary nonspherocytic haemolytic anaemia. It may also be important in the aetiology of the anaemia of lead intoxication because lead markedly inhibits P5N activity.Two enzymes involved in glutathione synthesis, y-glutamylcysteine synthetase (GC-S) and glutathione synthetase (GSH-S), were not included in the ICSH recommendations because the assay methods were complicated. GC-S deficiency and GSH-S deficiency both appear to be associated with moderate haemolytic anaemia and a marked decrease of red cell GSH (Beutler 1978) and thus the measurement of red cell GSH levels is useful as a screening procedure in diagnosing these two rare enzyme deficiencies. Diminished GSH stability is also a consistent finding in glucose 6-phosphate dehydrogenase (G6PD) deficiency, although the initial decreased GSH content of red cells is not marked.

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Seven Pyruvate Kinase Variants Characterized by the ICSH Recommended Methods

Cited by 50 publications

References 9 publications

cDNA cloning of human R-type pyruvate kinase and identification of a single amino acid substitution (Thr384----Met) affecting enzymatic stability in a pyruvate kinase variant (PK Tokyo) associated with hereditary hemolytic anemia.

cDNA cloning of human R-type pyruvate kinase and identification of a single amino acid substitution (Thr384----Met) affecting enzymatic stability in a pyruvate kinase variant (PK Tokyo) associated with hereditary hemolytic anemia.

The estimate of protein polymorphism in human populations: Lack of evidence for overestimation due to post-translational modification.

Recommended methods for an additional red cell enzyme (pyrimidine 5′-nucleotidase) assay and the determination of red cell adenosine-5′-triphosphate, 2,3-diphosphoglycerate and reduced glutathione

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