Behavior of caldesmon upon interaction of thin filaments with myosin subfragment 1 in ghost fibers.
Caldesmon is a component of smooth muscle thin filaments which inhibits their interaction with myosin. We have used polarized fluorescence technique to study the behavior of caldesmon during the interaction of myosin subfragment 1 (S1) with thin filaments reconstituted in rabbit skeletal muscle ghost fibers by incorporation of smooth muscle tropomyosin and caldesmon labeled with acrylodan at cysteine residue located in the C-terminal region. Significant changes in acrylodan fluorescence intensity upon addition of skeletal muscle S1 reflected substantial displacement of caldesmon from thin filaments, while alterations in the calculated fluorescence parameters indicated the simultaneous rearrangement of the remaining caldesmon fraction. The orientation of caldesmon in the S1-thin filament complex relative to the fiber axis changes by approximately 7 degrees and the mobility of the fluorescent probe by about 9%. The alterations in caldesmon orientation were proportional to the strength of S1 binding and diminished respectively upon addition of ADP and ADP-V(i). The changes in orientation of acrylodan-caldesmon evoked by the interaction of S1 with thin filaments were more pronounced than that in AEDANS-F-actin which suggests that the spatial arrangement of caldesmon in the complex is governed not only by F-actin but also by S1. The results may indicate that the changes in spatial arrangement of caldesmon are adjusted to the conformation of F-actin and S1 characteristic for particular steps of the ATP hydrolysis cycle.
Borovikov
Yurii S
YS
Laboratory of Mechanisms of Cell Motility, Institute of Cytology, Russian Academy of Sciences, 4 Tikhoretsky Avenue, St. Petersburg 194064, Russia.
Wrzosek
Antoni
A
Kulikova
Natalia
N
Vikhorev
Peter
P
Vikhoreva
Natalia
N
Dabrowska
Renata
R
D000199:Actins / Q000378:metabolism; D000244:Adenosine Diphosphate / Q000378:metabolism*; D000818:Animals; D002148:Calmodulin-Binding Proteins / Q000378:metabolism*; D002645:Chickens; D004372:Ducks; D005454:Fluorescence Polarization; D005456:Fluorescent Dyes; D005895:Gizzard, Avian / Q000737:chemistry; D018485:Muscle Fibers, Skeletal / Q000187:drug effects / Q000378:metabolism*; D018482:Muscle, Skeletal / Q000187:drug effects / Q000378:metabolism*; D015879:Myosin Subfragments / Q000378:metabolism*; D011487:Protein Conformation; D011817:Rabbits; D014335:Tropomyosin / Q000494:pharmacology
D000199:Actins; D002148:Calmodulin-Binding Proteins; D005456:Fluorescent Dyes; D015879:Myosin Subfragments; D014335:Tropomyosin; D000244:Adenosine Diphosphate