Actin-Tropomyosin Activation of Myosin Subfragment 1 ATPase and Thin Filament Cooperativity. The Role of Tropomyosin Flexibility and End-to-End Interactions

Lehrer, Sherwin S.; Golitsina, Nina L.; Geeves, Michael A.

doi:10.1021/bi971568w

Cited by 92 publications

(89 citation statements)

References 38 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Since the HMM effect usually levels off at a head/actin ratio between 3:7 and 5:7, the effect of HMM on tropomyosin must be more complex. A similar difference in cooperativity between skS1 and skHMM has been observed by others in the head-induced transition of smTm from off to on states (38,41).…”

Section: Resultssupporting

confidence: 83%

“…39 and references therein), and S1 or HMM molecules bind randomly (i.e. as a linear function of head concentration) to actin (38,40,41). We used the equation f m ϭ 1 Ϫ (1 Ϫ f b ) n (38), where f m is the fraction of tropomyosin molecules moved, f b is the fraction of actin monomers to which S1 or HMM is bound, and n, the cooperative unit, is the number of actin monomers affected, via the movement of tropomyosin, by the binding of each S1 or HMM molecule to the actin thin filament.…”

Section: Resultsmentioning

confidence: 99%

See 1 more Smart Citation

Phosphorylation of Smooth Muscle Myosin Heads Regulates the Head-induced Movement of Tropomyosin

Graceffa¹

2000

Journal of Biological Chemistry

View full text Add to dashboard Cite

It has been shown that skeletal and smooth muscle myosin heads binding to actin results in the movement of smooth muscle tropomyosin, as revealed by a change in fluorescence resonance energy transfer between a fluorescence donor on tropomyosin and an acceptor on actin (Graceffa, P. (1999) Biochemistry 38, 11984 -11992). In this work, tropomyosin movement was similarly monitored as a function of unphosphorylated and phosphorylated smooth muscle myosin double-headed fragment smHMM. In the absence of nucleotide and at low myosin head/actin ratios, only phosphorylated heads induced a change in energy transfer. In the presence of ADP, the effect of head phosphorylation was even more dramatic, in that at all levels of myosin head/actin, phosphorylation was necessary to affect energy transfer. It is proposed that the regulation of tropomyosin position on actin by phosphorylation of myosin heads plays a key role in the regulation of smooth muscle contraction. In contrast, actin-bound caldesmon was not moved by myosin heads at low head/actin ratios, as uncovered by fluorescence resonance energy transfer and disulfide cross-linking between caldesmon and actin. At higher head concentration caldesmon was dissociated from actin, consistent with the multiple binding model for the binding of caldesmon and myosin heads to actin (Chen,

show abstract

Section: Resultssupporting

confidence: 83%

Section: Resultsmentioning

confidence: 99%

Phosphorylation of Smooth Muscle Myosin Heads Regulates the Head-induced Movement of Tropomyosin

Graceffa¹

2000

Journal of Biological Chemistry

View full text Add to dashboard Cite

show abstract

“…Previous studies have used skTm, and there was no detectable binding for the Tm with an unmodified N-terminus (20). SmTm has a higher affinity for actin than skTm (29), and we show here differences of affinity of α-Tm to actin for native, and wild type with and without the AS extension. In the case of the αα homodimers, the AS extension increased the affinity by 100 fold and gave an affinity within a factor of 2 of the affinity of the native αβ-Tm.…”

Section: Discussionmentioning

confidence: 51%

Functional Homodimers and Heterodimers of Recombinant Smooth Muscle Tropomyosin

2006

Self Cite

View full text Add to dashboard Cite

Skeletal and smooth muscle tropomyosin (Tm) require acetylation of their N termini to bind strongly to actin. Tm containing an N-terminal alanine serine (AS) extension to mimic acetylation, has been widely used to increase binding. The current study investigates the ability of an N-terminal AS extension to mimic native acetylation for both αα and ββ smooth Tm homodimers. We show that : 1) ASα-Tm binds actin 100 fold tighter than α-Tm, and 2 fold tighter than native smooth αβ-Tm; 2) β-Tm requires an AS extension to bind actin; 3) ASβ-Tm binds actin 10 fold weaker than ASα-Tm. Tm is present in smooth muscle tissues as >95% heterodimer, therefore we studied the binding of recombinant αβ heterodimers with different AS extensions. This study shows that recombinant Tm requires an AS extension on both α and β chains to bind like native Tm and that the α chain contributes more to actin binding than the β chain. Once assembled onto an actin filament all smooth muscle Tms regulate S1 binding to actin Tm in the same way, irrespective of the presence of an AS extension.Tropomyosin (Tm) is an actin binding, α-helical, coiled coil protein dimer which binds along the length of actin filaments in both muscle and non-muscle cells, and thus cooperatively regulates the interaction of actin with myosin heads. Muscle cells contain Tm expressed via two genes Tm1 and Tm2 (α and β), and isoform diversity (smooth and skeletal) results from the alternative splicing of the two genes. Each monomer of the dimer coiled-coil contains 284 amino acid residues (1-3).In skeletal muscle tissue Tm is present predominantly as a mixture of αβ heterodimer and αα homodimer (4-6). ββ is less stable and is much less common in muscle tissues (5,6). Studies involving guanidinium hydrochloride dissociation of native homodimer chains and anion exchange chromatography have shown that smooth muscle contains roughly equal amounts of α and β chains, more than 95% of which is present as heterodimer (7-9). Thermal unfolding studies also show that the heterodimer is preferentially formed at temperatures below its unfolding transition (9). Preference of the heterodimer is not fully understood, however viscosity and thermal unfolding measurements indicate that the strength of the end-to-end interaction of the smooth αβ heterodimer is much higher than that of the αα and ββ homodimers which have about the same strength of end-to-end interaction (7,8,10). The strength of end-toend interactions in the heterodimer might affect the cooperative behaviour of the tropomyosin, and thus might provide a functional advantage for the predominance of heterodimer in native smooth muscle. The ability to be able to assemble in vitro heterodimers would be very valuable. Such ability could be used to monitor the behaviour of a single α or β chain on the dimer via

show abstract

“…ATPase assays were performed either by the colorimetric detection of phosphate described earlier (22) or by the continuous coupled enzyme assay system (23) EnzCheck (Molecular Probes).…”

Section: Isolation Of Proteins From Rabbit Skeletal Muscle-troponin Amentioning

confidence: 99%

A Modulatory Role for the Troponin T Tail Domain in Thin Filament Regulation

Maytum¹,

Geeves²,

Lehrer³

2002

Journal of Biological Chemistry

Self Cite

View full text Add to dashboard Cite

In striated muscle the force generating acto-myosin interaction is sterically regulated by the thin filament proteins tropomyosin and troponin (Tn), with the position of tropomyosin modulated by calcium binding to troponin. Troponin itself consists of three subunits, TnI, TnC, and TnT, widely characterized as being responsible for separate aspects of the regulatory process. TnI, the inhibitory unit is released from actin upon calcium binding to TnC, while TnT performs a structural role forming a globular head region with the regulatory TnITnC complex with a tail anchoring it within the thin filament. We have examined the properties of TnT and the TnT 1 tail fragment (residues 1-158) upon reconstituted actin-tropomyosin filaments. Their regulatory effects have been characterized in both myosin S1 ATPase and S1 kinetic and equilibrium binding experiments. We show that both inhibit the actin-tropomyosin-activated S1 ATPase with TnT 1 producing a greater inhibitory effect. The S1 binding data show that this inhibition is not caused by the formation of the blocked B-state but by significant stabilization of the closed C-state with a 10-fold reduction in the C-to M-state equilibrium, K T , for TnT 1 . This suggests TnT has a modulatory as well as structural role, providing an explanation for its large number of alternative isoforms.

show abstract

Actin-Tropomyosin Activation of Myosin Subfragment 1 ATPase and Thin Filament Cooperativity. The Role of Tropomyosin Flexibility and End-to-End Interactions

Cited by 92 publications

References 38 publications

Phosphorylation of Smooth Muscle Myosin Heads Regulates the Head-induced Movement of Tropomyosin

Phosphorylation of Smooth Muscle Myosin Heads Regulates the Head-induced Movement of Tropomyosin

Functional Homodimers and Heterodimers of Recombinant Smooth Muscle Tropomyosin

A Modulatory Role for the Troponin T Tail Domain in Thin Filament Regulation

Contact Info

Product

Resources

About