Studies on cell-free metabolism: Ethanol production by a yeast glycolytic system reconstituted from purified enzymes

Welch, Peta; Scopes, Robert K.

doi:10.1016/0168-1656(85)90029-x

Cited by 91 publications

(74 citation statements)

References 24 publications

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“…NATURE COMMUNICATIONS | DOI: 10.1038/ncomms5113 ARTICLE biochemistry system employing glycolysis has been demonstrated previously 5 . Building more complex compounds from acetyl-CoA such as fatty acids, polyketides and other isoprenoids will also require the incorporation and recycling of ATP.…”

Section: Discussionmentioning

confidence: 98%

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A synthetic biochemistry molecular purge valve module that maintains redox balance

2014

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Section: Discussionmentioning

confidence: 98%

“…Building single, dedicated pathways in vitro can eliminate side reactions that occur in the cell, so that nearly 100% yields and fast reaction times are possible [3][4][5] . In vitro biochemical systems also allow for more precise control over optimization and product toxicity problems can be more easily diagnosed and mitigated 6,7 .…”

mentioning

confidence: 99%

A synthetic biochemistry molecular purge valve module that maintains redox balance

2014

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“…In Z. mobilis we find that the ratios between the enzymes do differ significantly from those reported for eukaryotic sources. Glyceraldehyde-phosphate dehydrogenase is not present at such a high activity as in yeast (Welch & Scopes, 1985;, although the glycolytic rate in Z. mobilis is greater than in yeasts (Lee et al, 1979). The reason may be found by considering the preceding reactions in the two species; the virtual absence of triosephosphate isomerase in Z. mobilis means that the 3-deoxy-2-oxo-6-phosphogluconate aldolase is able to provide a relatively high concentration of glyceraldehyde phosphate.…”

Section: Discussionmentioning

confidence: 99%

Isolation and properties of the glycolytic enzymes from Zymomonas mobilis. The five enzymes from glyceraldehyde-3-phosphate dehydrogenase through to pyruvate kinase

Pawluk¹,

Scopes²,

Griffiths-Smith³

1986

Biochemical Journal

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The five glycolytic enzymes glyceraldehyde-3-phosphate dehydrogenase, phosphoglycerate kinase, phosphoglycerate mutase, enolase and pyruvate kinase were each purified from extracts of Zymomonas mobilis cells, by using dye-ligand chromatography as the principal step. Two procedures, producing three and two of the enzymes respectively, are described in detail. Z. mobilis glyceraldehyde-phosphate dehydrogenase was found to be similar in most respects to the enzyme from other sources, except for having a slightly larger subunit size. Phosphoglycerate kinase has properties typical for this enzyme; however, it did not show the sulphate activation effects characteristic of this enzyme from most other sources. Phosphoglycerate mutase is a dimer, partially independent of 2,3-bisphosphoglycerate, and has a high specific activity. Enolase was found to be octameric; otherwise its properties were very similar to those of the yeast enzyme. Pyruvate kinase is unusual in being dimeric, and not requiring K+ for activity. It is not allosterically activated by sugar phosphates, having a high activity in the absence of any effectors. Some quantitative differences in the relative amounts of these enzymes, compared with eukaryotic species, are ascribed to the fact that Z. mobilis utilizes the Entner-Doudoroff pathway rather than the more common Embden-Meyerhoff glycolytic route.

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“…The activity of the enzyme in Z. mobilis, at around 600 p.mol min1 g (wet weight)1-(1), is higher than in any other source; even yeasts do not have such high levels in their cytoplasm (32). The enzyme constitutes over 5% of the readily soluble protein in Z. mobilis grown under normal conditions with glucose as substrate, at 30 to 32°C, pH 5 to 6.…”

mentioning

confidence: 94%

“…Enzyme activity was lost if the extract was exposed to a pH of >7.0. Recombinant E. coli cells were extracted by a similar procedure, with the addition of vigorous vortexing in the presence of 50-to 150-,um glass beads (Sigma) (32).…”

Section: Methodsmentioning

confidence: 99%

Pyruvate decarboxylase of Zymomonas mobilis: isolation, properties, and genetic expression in Escherichia coli

Neale¹,

Scopes²,

Wettenhall³

et al. 1987

J Bacteriol

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Pyruvate decarboxylase (EC 4.1.1.1) from Zymomonas mobilis purified to homogeneity by using dye-ligand and ion-exchange chromatography. Antibodies produced against the enzyme and the amino-terminal sequence obtained for the pure enzyme were used to select and confirm the identity of a genomic clone encoding the enzyme selected from a genomic library of Z. mobilis DNA cloned into pUC9. The genomic fragment encoding the enzyme expressed high levels of pyruvate decarboxylase in Escherichia coli. Possible RNA polymerase and ribosome-binding sites have been identified in the 5'-untranslated region of the pyruvate decarboxylase gene.

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Studies on cell-free metabolism: Ethanol production by a yeast glycolytic system reconstituted from purified enzymes

Cited by 91 publications

References 24 publications

A synthetic biochemistry molecular purge valve module that maintains redox balance

A synthetic biochemistry molecular purge valve module that maintains redox balance

Isolation and properties of the glycolytic enzymes from Zymomonas mobilis. The five enzymes from glyceraldehyde-3-phosphate dehydrogenase through to pyruvate kinase

Pyruvate decarboxylase of Zymomonas mobilis: isolation, properties, and genetic expression in Escherichia coli

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