Caldesmon reduces the apparent rate of binding of myosin S1 to actin-tropomyosin.
Equilibrium measurements of the rate of binding of caldesmon and myosin S1 to actin-tropomyosin from different laboratories have yielded different results and have led to different models of caldesmon function. An alternate approach to answering these questions is to study the kinetics of binding of both caldesmon and S1 to actin. We observed that caldesmon decreased the rate of binding of S1 to actin in a concentration-dependent manner. The inhibition of the rate of S1 binding was enhanced by tropomyosin, but the effect of tropomyosin on the binding was small. Premixing actin with S1 reduced the amplitude (extent) of caldesmon binding in proportion to the fraction of actin that contained bound S1, but the rate of binding of caldesmon to free sites was not greatly altered. No evidence for a stable caldesmon-actin-tropomyosin-S1 complex was observed, although S1 did apparently bind to gaps between caldesmon molecules. These results indicate that experiments involving caldesmon, actin, tropomyosin, and myosin are inherently complex. When the concentration of either S1 or caldesmon is varied, the amount of the other component bound to actin-tropomyosin cannot be assumed to remain fixed. The results are not readily explained by a mechanism in which caldesmon acts only by stabilizing an inactive state of actin-tropomyosin. The results support regulatory mechanisms that involve changes in the actin-S1 interaction.
Sen
A
A
Department of Biochemistry, Brody School of Medicine at East Carolina University, Greenville, North Carolina 27858-4354, USA.
Chen
Y D
YD
Yan
B
B
Chalovich
J M
JM
D000199:Actins / Q000378:metabolism*; D000818:Animals; D002148:Calmodulin-Binding Proteins / Q000378:metabolism*; D019793:Fluorescein / Q000378:metabolism; D005452:Fluoresceins / Q000378:metabolism; D005456:Fluorescent Dyes / Q000378:metabolism; D007460:Iodoacetamide / Q000031:analogs & derivatives* / Q000378:metabolism; D007700:Kinetics; D008027:Light; D015879:Myosin Subfragments / Q000037:antagonists & inhibitors* / Q000378:metabolism*; D010069:Oxadiazoles / Q000378:metabolism; D011485:Protein Binding; D011721:Pyrenes / Q000378:metabolism; D011817:Rabbits; D012459:Salicylates / Q000378:metabolism; D012542:Scattering, Radiation; D013050:Spectrometry, Fluorescence; D014335:Tropomyosin / Q000378:metabolism*; D014422:Turkeys
D000199:Actins; D002148:Calmodulin-Binding Proteins; D005452:Fluoresceins; D005456:Fluorescent Dyes; D015879:Myosin Subfragments; D010069:Oxadiazoles; D011721:Pyrenes; D012459:Salicylates; D014335:Tropomyosin; C005487:4-iodoacetamidosalicylic acid; C015296:5-iodoacetamidofluorescein; C035573:4-(N-(iodoacetoxy)ethyl-N-methyl)amino-7-nitrobenz-2-oxa-1,3-diazole; C030984:pyrene; D019793:Fluorescein; D007460:Iodoacetamide