Peptide structures of pyruvate kinase isozymes: 2. Origins of types M1 and M2 isozymes suggested from species-variations in their peptide maps

doi:10.1016/0167-4838(82)90072-3

Biochimica et Biophysica Acta (BBA) - Protein Structure and Molecular Enzymology

Volume 704, Issue 3, 24 June 1982, Pages 494-502

Biochimica et Biophy...

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Abstract

Pyruvate kinase (ATP: pyruvate 2-O-phosphotransferase, EC 2.7.1.40) isozymes were purified: type M₁ and M₂ isozymes from rat, mouse and rabbit, type M₁ from bull frog and type L from rat and mouse. The amino acid compositions of type M₁ and M₂ isozymes from various species were very similar and those of type L isozymes from rat and mouse were also very similar. The NH₂-terminals of type M₁ and M₂ from rat and mouse were (Pro-Lys-Pro-), but those of the other enzymes appeared to be blocked. The COOH-terminals of type M₁ and M₂ from rat, mouse and rabbit were (-Val-Pro). Comparison of tryptic peptide maps of type M₁ and M₂ from rat, mouse and rabbit indicated that type M₁ and M₂ could not be produced by post-synthetic modification, but could be encoded by different type-specific mRNAs. Comparative studies on type MI and type M 2 of different species showed that type M₁-specific peptide spots were highly variable, whereas type M₂- specific peptide spots were highly conserved. The type L isozyme showed marked species variation, indicating that this differentiated isozyme evolved more rapidly than type M₂, which may be a prototype or undifferentiated isozyme.

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Cited by (15)

Phylogeny congruence analysis and isozyme classification: The pyruvate kinase system
1989, Journal of Theoretical Biology
As the isozymes of pyruvate kinase (PK) are best known in rats, the characteristics of the rat isozymes are generally used to classify the PK isozymes in other species. Given the discrepancies generated by this classification by analogy, we evaluated a classification using a phylogeny congruence analysis of the compositional relatedness of vertebrate PK's. While our phylogenetic analysis confirmed the well established separation of the L and R isozymes from the K and M isozymes, its power became most evident in the identification of non-orthologous (or variant) forms of PK. Our analysis emphasized the uniqueness of chicken liver PK which cannot be classified either as a K or an L isozyme, confirmed that tumors express a variety of forms of PK, and indicated that lungs systematically express PK's which are not orthologous with PK's from other tissues. The determination of orthology by the phylogeny congruence analysis assumes that the structural data from different sources are subject to similar methodological error. However, we cannot reject the possibility that an apparent lack of orthology be due to artifacts during purification and analysis.
Pyruvate Kinase
1987, Enzymes
The chapter discusses the regulatory phosphorylation of pyruvate kinase type L by cAMP-dependent protein kinase. It also briefly discusses the role of phosphorylation of liver pyruvate kinase in the regulation of glycolysis and gluconeogenesis. The phosphorylation of liver pyruvate kinase alters the kinetic behavior of the enzyme, including the influence of allosteric effectors on the enzyme. The dephosphorylation of pyruvate kinase is mainly accounted for by protein phosphatase-2A. For the phosphorylation of the enzyme by cAMP-dependent protein kinase, the structural requirements reside mainly in the amino acid sequence of the phosphorylatable site. As its counterpart in the liver, kidney pyruvate kinase—type L—is phosphorylated on serine residues upon incubation with ATP and cAMP-dependent protein kinase. The phosphorylation of the pyruvate kinase and the changes of its activity are reversed by phosphoprotein phosphatase. In vivo and in isolated hepatocytes, the enzyme is subject to the same functional changes in the presence of glucagon or cAMP as the purified enzyme when it is incubated with cAMP-dependent protein kinase and ATP. The degree of phosphorylation appears to correlate fairly well with the changes in the kinetic properties of the enzyme in vitro and with the hormonal response and cAMP level in intact cells.
Pyruvate kinase and total protein are regulated differently during growth of P-815 mastocytoma cells
1986, Archives of Biochemistry and Biophysics
Total protein content of P-815 mastocytoma cells decreases and then increases in response to initiation of a new growth cycle. As the level increases, the rate of synthesis declines. Both events occur prior to any decrease in the rate of cellular proliferation. These temporal relationships indicate that the rate of protein synthesis reflects the intracellular concentration of protein rather than the cellular growth rate, as has been hypothesized. Pyruvate kinase protein metabolism differs from that of total protein in three ways: (a) accumulation does not stop, (b) the rate of synthesis does not decrease, and (c) only the rate of pyruvate kinase degradation is altered by a factor present in conditioned media. These observations suggest that there are specific mechanisms regulating pyruvate kinase at a post-transcriptional level.
Purification of pyruvate kinase isoenzymes type M<inf>1</inf> and M<inf>2</inf> from dog (Canis familiaris) and comparison of their properties with those from chicken and rat
1986, Comparative Biochemistry and Physiology -- Part B: Biochemistry and
- 1.
  1. Pyruvate kinase isoenzymes type M₁ and M₂ from dog muscle, lung and tumor have been isolated.
- 2.
  2. The K_0.5 for phosphoenol pyruvate have been determined to be0.04 mM for dog muscle type M₁, 0.24 mM for lung type M₂ and 0.28 mM for tumor type M₂ isoenzymes.
- 3.
  3. The activator constant k_a of l-serine is 240 nM from lung and 70 nM from tumor isoenzyme.
- 4.
  4. Consistent with the assumption of a special form of pyruvate kinase M₂ in dog tumor cells different isoelectric points and amino acid compositions have been found for the isoenzymes of lung and tumor.
The purification and kinetic characterization of eel white muscle pyruvate kinase
1985, Comparative Biochemistry and Physiology -- Part B: Biochemistry and
- 1.
  1. A stable, homogeneous preparation of pyruvate kinase from white muscle of the American eel, Anguilla rostrata with a specific activity of 350 units/mg has been obtained.
- 2.
  2. The enzyme has a pH optimum in the range 6.3–6.5 and requires Mg²⁺ and K⁺ for maximum activity.
- 3.
  3. Eel muscle pyruvate kinase exhibits slight co-operativity in the binding of the substrate phosphoenol-pyruvate. It is activated by fructose-1,6-biphosphate in a pH dependent manner and is inhibited by both alanine and phenylalanine. These properties are very similar to the properties of the mammalian M₂ isozyme.
Peptide structures of pyruvate kinase isozymes. 1. Comparison of the four pyruvate kinase isozymes of the rat
1982, Biochimica et Biophysica Acta (BBA)/Protein Structure and Molecular
The four isozymes of rat pyruvate kinase (ATP:pyruvate 2-O-phosphotransferase, EC 2.7.1.40) were purified by immunoadsorbent column chromatography or affinity elution chromatography, and their peptide structures were compared. Their apparent subunit molecular weights, estimated by SDS-polyacrylamide gel electrophoresis, were 59 000 (type Ml), 60 000 (type M₂), 58500 (type L) and 62 000 (type R). Comparison of the peptide patterns produced by CNBr cleavage and trypsin digestion showed clear differences between the peptide structures of type M₁ and M₂, and type L and R, respectively. It also showed remarkable structural homology between them. Terminal analyses showed that type M₁ and M₂ have the same NH₂-terminal sequence (Pro-Lys-Pro-) and the same COOH-terminal sequence (-Val-Pro). Type L and R have no free NH2-terminal residue, but probably have the same COOH-terminal sequence, (-Ser-Val) or (-Val-Ser). Findings on types M₁ and M₂ exclude the possibUity of their post-synthetic precursor-product relationship; they are probably produced by specific mRNAs transcribed from a single gene through rather different processings, or transcribed from the separate, but closely related genes. The structural differences between types L and R may be in the NH2-terminal region, but the NH₂-terminals are blocked, lmmunoadsorbent purification gave no evidence for their precursor-product relationship, but showed tissue-specific production of types R and L. The evidence supports the idea proposed by Marie et al. (Nature 1981, 292, 70–72) that types L and R are produced by specific mRNAs transcribed from a single gene through different processings.

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^∗: Present address: Deaprtment of Medical Biochemistry, School of Medicine, Ehime University, Shigenobu, Ehime 791-02, Japan.

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Peptide structures of pyruvate kinase isozymes: 2. Origins of types M1 and M2 isozymes suggested from species-variations in their peptide maps

Abstract

Biochim. Biophys. Acta

J. Biol. Chem.

Int. J. Biochem.

Comp. Biochem. Physiol.

Biochem. Biophys. Res. Commun.

J. Biol. Chem.

Biochem. Biophys. Res. Commun.

Biochim. Biophys. Acta

J. Biol. Chem.

Peptide structures of pyruvate kinase isozymes: 2. Origins of types M₁ and M₂ isozymes suggested from species-variations in their peptide maps