Comparison of ability of Mg and Mn to activate the key enzymes of glycolysis

Wimhurst, Janet M.; Manchester, Keith

doi:10.1016/0014-5793(72)80650-1

Cited by 27 publications

(8 citation statements)

References 22 publications

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“…This result is consistent with Bygrave's (1966) finding. We have 1975 also shown, by calculation, that Cm decreases as At or Bt decreases, in agreement with findings of Wimhurst & Manchester (1972). It has already been noted that the maxima in the plots v = f(C) are particularly broad when C is Mg2+ and when either a and B or A and ,B are kept constant ( Fig.…”

Section: Kinetic Constantssupporting

confidence: 87%

“…Turning now to more qualitative aspects of our results, note that the differences in the influence of the bivalent cations, referred to above, have been observed in solutions with comparable ionic strengths. We can therefore, in agreement with Wimhurst & Manchester (1972), set aside ionic-strength effects as the possible cause of the inhibitions observed at high concentrations of bivalent cations.…”

Section: Kinetic Constantssupporting

confidence: 86%

“…However, when the total concentrations of ADP and phosphoenolpyruvate are kept constant, the activation that is brought about by the bivalent cation is succeeded by inhibition as its concentration is increased (Ainsworth & Macfarlane, 1973; Bygrave, 1966). It has also been observed that the concentration of bivalent cation required for maximum activity depends both oD the identity of the cation and on the concentrations of ADP and phosphoenolpyruvate used (Wimhurst & Manchester, 1972).…”

mentioning

confidence: 99%

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Activation and inhibition of rabbit muscle pyruvate kinase by transition-metal ions

Ainsworth¹,

MacFarlane²

1975

Biochemical Journal

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The paper reports a comparative study of the effects of Mn2+, Ni2+ and Co2+ on the reaction of ADP with phosphoenolypyruvate when catalysed by K+-activated rabbit muscle pyruvate kinase. The activation and subsequent inhibition that occurs as the bivalent ion concentration is increased is taken as evidence that the substrates of the enzyme are phosphoenolypyruvate, uncomplexed ADP and free bivalent cation. Kinetic constants for the binding of the bivalent cation to the enzyme are reported.

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Section: Kinetic Constantssupporting

confidence: 87%

Section: Kinetic Constantssupporting

confidence: 86%

mentioning

confidence: 99%

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Activation and inhibition of rabbit muscle pyruvate kinase by transition-metal ions

Ainsworth¹,

MacFarlane²

1975

Biochemical Journal

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“…Studies in vitro have shown that presence of Mn +2 ions can impact cleavage activity of Cas9 and Cas12a without a gRNA present (44). Our injection buffer includes a functionally similar ion, Mg +2 , which has been shown to compete with Mn +2 and activate common enzymes (55). Therefore, we investigated the effect of Cas9 in genomic DNA of larvae solely injected with the nuclease, either as an enzyme or mRNA.…”

Section: Discussionmentioning

confidence: 99%

Evaluation of CRISPR gene-editing tools in zebrafish

Uribe-Salazar

Sekar

Kaya

et al. 2020

Preprint

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Zebrafish have practical features that make them a useful model for higher-throughput tests of gene function using CRISPR/Cas9 editing to create ‘knockout’ models. Due to the large number of available tools to design CRISPR assays and diversity of theories/model systems they were originally built on, we sought to systematically compare computational and empirical approaches for predicting gene-editing efficacy in zebrafish. We subjected zebrafish embryos to CRISPR/Cas9 with 50 different guide RNAs (gRNAs) targeting 14 genes and assayed individual editing efficiencies. We compared our experimental in vivo efficiencies in mosaic G0 embryos with those predicted by seven commonly used gRNA design tools and found large discrepancies between methods. Assessing off-target mutations (predicted in silico and in vitro) found that the majority of tested loci had low in vivo frequencies (<1%). Moreover, understanding that recent segmental duplications in the zebrafish genome could exacerbate CRISPR targeting of individual genes, we cataloged these loci and have made them available as a resource. Lastly, we assessed the transcriptome of negative ‘mock’ control CRISPR larvae injected with Cas9 enzyme or mRNA with no gRNA using RNA-seq and identified differentially expressed genes with high variability between injections. Using these same data, we discovered on average ~60 putative somatic mosaic frameshift mutations impacting genes per pool of injected larvae, potentially due to background cutting of DNA with Cas9 in the absence of gRNA. To verify this previously unreported phenomenon in zebrafish, we validated seven of twelve genes tested carrying low frequency mosaic somatic mutations in the genomes of a separate batch of injected larvae. These results suggest that negative control embryos may carry mutations within genes leading to spurious phenotypes. Overall, our results provide a valuable resource for the zebrafish community for the design and execution of CRISPR/Cas9 experiments.

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“…Under these conditions ATP 4-increased the allosteric inhibition by MgATP 2-. The requirement for free Mg z+ in amounts equal to the substrate MglTP 2-concentration suggests that Mg 2+ has at least two roles in the reaction mechanism: (a) it forms a complex with ITP 4-to form MglTP 2-which is the substrate for the enzyme, and (b) it forms a complex with the enzyme to activate the enzymic reaction [12][13][14][15][16]. In contrast to the enzyme from yeast [1], platelet phosphofructokinase is inhibited by free ATP 4-, since these ions increase the allosteric inhibition by the MgATP zcomplex.…”

Section: Discussionmentioning

confidence: 99%