Nanosecond pulse fluorometry in polarized light of dansyl‐L‐cysteine linked to a unique SH group of F‐actin; The influence of regulatory proteins and myosin moiety

Wahl, Philippe; Mihashi, Koshin; Auchet, Jean-Claude

doi:10.1016/0014-5793(75)80443-1

Cited by 22 publications

(14 citation statements)

References 14 publications

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“…For G-actin, the observed correlation times are roughly consistent with the size and shape of the actin monomer. For F-actin, both labels yielded rotational correlation times of the order of 500 ns, in agreement with [4,5]. After due consideration of all the other pertinent findings, these new results indicate that the observed correlation times of about 500 ns correspond to flexibility of the covalent fluorescence labels at their attachment sites.…”

supporting

confidence: 64%

“…As pointed out [4] , it is also possible that the protein segment containing the labeling site is flexible. F-a&in was spin-labeled with a maleimide derivative [6], probably at the same penultimate cysteine 373.…”

Section: Anisotropy Decay For Labeled F-actinmentioning

confidence: 99%

“…The covalent probes 1 &IAEDANS (this work), didansyl cystine [4], pyrene maleimide [5 ] and maleimide spin label [6] all possess at least 3 bonds that have complete torsional freedom. The presence of label ~e~b~ity had been suggested for eosin iso~o~anate-labeled ovalbumin [23 1, and for dansyl chloride-labeled membrane fragments [24].…”

Section: Anisotropy Decay For Labeled F-actinmentioning

confidence: 99%

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Nanosecond fluorescence depolarization studies on actin labeled with 1,5‐IAEDANS and dansyl chloride

Tao

1978

FEBS Letters

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supporting

confidence: 64%

Section: Anisotropy Decay For Labeled F-actinmentioning

confidence: 99%

Section: Anisotropy Decay For Labeled F-actinmentioning

confidence: 99%

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Nanosecond fluorescence depolarization studies on actin labeled with 1,5‐IAEDANS and dansyl chloride

Tao

1978

FEBS Letters

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“…To try to answer that question, we recently studied the anisotropy decay of fluorescent chromophores, covalently linked to the residue cysteine-373 of actin complexed with troponin and tropomyosin [7,8]. It has been found that the correlation time of these chromophores decreased in the presence of micromolar concentrations of Ca2+ and that, under this condition, the correlation time was smaller than the one…”

mentioning

confidence: 99%

“…F actin forms complexes with tropomyosin and troponin in which the arrangement of these molecules is the same as in the thin muscle filaments. Studies of these reconstituted filaments suggest, in contradistinction to the steric blocking model, that changes in actin conformation could be an important factor in the regulation of the muscular contraction [4-61.To try to answer that question, we recently studied the anisotropy decay of fluorescent chromophores, covalently linked to the residue cysteine-373 of actin complexed with troponin and tropomyosin [7,8]. It has been found that the correlation time of these chromophores decreased in the presence of micromolar concentrations of Ca2+ and that, under this condition, the correlation time was smaller than the one…”

mentioning

confidence: 99%

Anisotropy Decay of Labelled Actin

Ikkai

Wahl

Auchet

1979

European Journal of Biochemistry

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G actin, labelled presumably on cysteine-373 with the fluorescent chromophore N-iodoacetyl-N'-(5 sulfo-1 -naphthyl)-ethylenediamine and purified by Sephacryl S-200 gel chromatography, migrated in one band on polyacrylamide gel electrophoresis and had the same polymerizability as unlabelled purified G actin. Anisotropy decays of labelled actin solutions have been studied at different ionic strengths and protein concentrations.It was found that these anisotropy decays could be fitted by a sum' of two exponential functions. Under low ionic strength or below the critical concentrations the longer correlation time (45 ns at 3.5 " C ) was independent of protein concentration and ionic strength. Above the critical concentration, the longer correlation time increased with ionic strength and protein concentration. In order to take into account that, under these conditions, the solutions contained a mixture of F and G actin at the critical concentration, the anisotropy decays were analysed as a sum of three exponential functions in which the longest correlation time characterized F actin. Since F actin correlation time also depended on actin concentration, an analysis with a sum of four exponential functions was performed, in which two fixed correlation times (100 ns and 900 ns at 3.5 "C) were introduced in order to characterize the F actin motions. The longer of these correlation times was attributed to regions where two actin filaments interact side by side, while the shorter one was attributed to filament regions free from intermolecular interactions.The small value of the free F actin correlation time indicates that the protomer peptide chain is very flexible around its C terminus, probably involving the motion of a molecular lobe. This flexibility might be an important factor in the interaction of actin with myosin during the muscular contraction.During muscular contraction, the globular heads of the myosin molecules which constitute the thick filament, interact cyclically with the actin protomers in the thin filaments [l].The thin filament also includes troponin and tropomyosin, the regulatory proteins which control the myosin-actin interaction, according to the calcium ion concentration in the cell [2]. Tropomyosin has been shown to lie along the grooves of the double-helix structure formed by the actin protomers in the filament. According to X-ray diffraction experiments, its position differs in the relaxed and the contracting states of the muscle [3]. The regulatory mechanism associated with this motion of tropomyosin has been explained by the steric blocking model [4], in which actin does not play an active role.Actin can be extracted from muscles with a lowionic-strength solvent in which this protein is dispersed as monomer molecules (G actin). If one increases the ionic strength, actin polymerizes in helical aggregates (F actin) [4]. F actin forms complexes with tropomyosin and troponin in which the arrangement of these molecules is the same as in the thin muscle filaments. Studies of these reconstituted filam...

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The Study of Cytoskeletal Protein Interactions by Fluorescence Probe Techniques

Blatt

Sawyer

1988

Subcellular Biochemistry

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Nanosecond pulse fluorometry in polarized light of dansyl‐L‐cysteine linked to a unique SH group of F‐actin; The influence of regulatory proteins and myosin moiety

Cited by 22 publications

References 14 publications

Nanosecond fluorescence depolarization studies on actin labeled with 1,5‐IAEDANS and dansyl chloride

Nanosecond fluorescence depolarization studies on actin labeled with 1,5‐IAEDANS and dansyl chloride

Anisotropy Decay of Labelled Actin

The Study of Cytoskeletal Protein Interactions by Fluorescence Probe Techniques

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