Fluorescence detection of increased local flexibility induced by coenzyme A in succinyl-CoA synthetase from Escherichia coli

Prasad, Anand; Nishimura, Jonathan S.; Horowitz, Paul M.

doi:10.1021/bi00264a006

Cited by 3 publications

(4 citation statements)

References 29 publications

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“…These results suggest that there are tryptophan residues at or near the CoA binding site and emphasize the necessity for the specific binding of CoA. Our earlier studies have shown that the binding of CoA does not bring about any large conformational changes in SCS (Prasad et al, 1982).…”

Section: Resultssupporting

confidence: 50%

“…This has been attributed to changes in SCS structure as a result of CoA binding. Our recent investigation using the dansyl derivative of SCS has shown that there are flexible segments in this protein and their flexibility is altered by the binding of CoA (Prasad et al, 1982). One of the limitations of this approach was the possibility of an alteration in the protein structure as a result of dansylation.…”

Section: Resultsmentioning

confidence: 99%

“…Implicit in this is the concept that binding of substrates brings about conformational changes in the region of the active site. Recently, we have been able to show through fluorescence measurements on dansyl-labeled enzyme that the regions of the molecules bearing a dansyl group, presumably close to the active site, showed greatly increased flexibility when CoA was present (Prasad et al, 1982). Because both subunits contained the dansyl label, it was not possible to determine where, a subunit, ß subunit, or both, the flexible regions lay.…”

mentioning

confidence: 99%

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A study of the quenching of the intrinsic fluorescence of succinyl-CoA synthetase from Escherichia coli by acrylamide, iodide, and coenzyme A

Prasad¹,

Nishimura²,

Horowitz³

1983

Biochemistry

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Escherichia coli succinyl-CoA synthetase (SCS) contains three tryptophan residues per mole of alpha beta dimer, and all of them are on the beta subunit. SCS shows an emission maximum at 335 nm which is shifted to 350 nm upon denaturation by urea or guanidine hydrochloride. Acrylamide is able to quench the tryptophan fluorescence in SCS by static and dynamic mechanisms. Substrates give protection against quenching by acrylamide. Binding of ATP to the alpha subunit which has no tryptophans gives as large an effect on the quenching by acrylamide as the binding of coenzyme A (CoA) to the beta subunit. Addition of CoA eliminates the curvature observed in Stern-Volmer plots for acrylamide quenching obtained by lifetime measurements. Potassium iodide does not quench the SCS fluorescence in the presence of CoA. These results suggest that there are heterogeneously emitting tryptophan residues in SCS that are located at the alpha beta subunit contact region close to the CoA binding site. Hence, the tryptophan residues can act as intrinsic reporters of events taking place at the active site of this enzyme. Further, the present results support models for SCS that put the active site at the alpha beta subunit contact region.

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Section: Resultssupporting

confidence: 50%

Section: Resultsmentioning

confidence: 99%

mentioning

confidence: 99%

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A study of the quenching of the intrinsic fluorescence of succinyl-CoA synthetase from Escherichia coli by acrylamide, iodide, and coenzyme A

Prasad¹,

Nishimura²,

Horowitz³

1983

Biochemistry

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“…This observation is consistent with a change in conformation of the protein upon binding of CoA. It was also observed that binding of CoA induced changes in the flexibility of the protein in regions labeled with dansyl chloride (85). (98,99).…”

Section: Phosphorytation Sitesupporting

confidence: 85%

Succinyl‐CoA Synthetase Structure‐Function Relationships and Other Considerations

Nishimura

1986

Advances in Enzymology - And Related Areas of Molecular Biology

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Strain energy in enzyme–substrate binding: An energetic insight into the flexibility versus rigidity of enzyme active site

Guo

Huang

et al. 2012

Computational and Theoretical Chemistry

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Fluorescence detection of increased local flexibility induced by coenzyme A in succinyl-CoA synthetase from Escherichia coli

Cited by 3 publications

References 29 publications

A study of the quenching of the intrinsic fluorescence of succinyl-CoA synthetase from Escherichia coli by acrylamide, iodide, and coenzyme A

A study of the quenching of the intrinsic fluorescence of succinyl-CoA synthetase from Escherichia coli by acrylamide, iodide, and coenzyme A

Succinyl‐CoA Synthetase Structure‐Function Relationships and Other Considerations

Strain energy in enzyme–substrate binding: An energetic insight into the flexibility versus rigidity of enzyme active site

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