TY - JOUR
T1 - Interaction of the heavy and light chains of cardiac myosin subfragment- 1 with F-actin
AU - Andreev, O. A.
AU - Borejdo, J.
PY - 1997
Y1 - 1997
N2 - The interaction of the heavy chain (HC) and the light chain (cdLC1) of cardiac S1 (cdS1) with F-actin was studied by cross-linking, Western blotting, and fluorescence polarization methods. Incorporation of cdLC1 in cross-linked products was examined by Western blots using the primary antibody against 71-74 residues of cdLC1. Cross-linking with zero-length, water-soluble reagent yielded three products with apparent molecular masses of 150, 160, and 210 kD. Like in the case of cross-linking of skeletal S1 with actin, these complexes included only HC of S1 and actin. The composition of the products were as follows: 150 kD, one HC of S1 cross-linked through a primary site (on the C-terminal of the 20-kD fragment) to the N-terminus of actin; 160 kD, one HC of S1 cross-linked through a secondary site (on the 50 kD fragment) to the N-terminus of actin; and 210 kD, one HC of S1 cross- linked through primary and secondary sites to two actins. Four additional products with apparent molecular masses of 66, 120, 185, and 235 kD contained cdLC1 and were identified as cdLC1+actin, cdLC1+HC(s1), cdLC1+actin+HC(s1), and cdLC1+two actins+HC(s1), respectively. The same products were observed when cross-linking was performed in cardiac myofibrils incubated with cdS1. The production of cross-linked complexes of the heavy and light chain with actin decreased with an increase in the molar ratio of cdS1:actin. To test whether the orientation of myosin heads depended on a degree of occupation of thin filaments, myofibrils were irrigated with varying concentrations of cdS1. Fluorescence polarization measurements of cdS1 bound to individual t- bands revealed that the orientation depended on the concentration.
AB - The interaction of the heavy chain (HC) and the light chain (cdLC1) of cardiac S1 (cdS1) with F-actin was studied by cross-linking, Western blotting, and fluorescence polarization methods. Incorporation of cdLC1 in cross-linked products was examined by Western blots using the primary antibody against 71-74 residues of cdLC1. Cross-linking with zero-length, water-soluble reagent yielded three products with apparent molecular masses of 150, 160, and 210 kD. Like in the case of cross-linking of skeletal S1 with actin, these complexes included only HC of S1 and actin. The composition of the products were as follows: 150 kD, one HC of S1 cross-linked through a primary site (on the C-terminal of the 20-kD fragment) to the N-terminus of actin; 160 kD, one HC of S1 cross-linked through a secondary site (on the 50 kD fragment) to the N-terminus of actin; and 210 kD, one HC of S1 cross- linked through primary and secondary sites to two actins. Four additional products with apparent molecular masses of 66, 120, 185, and 235 kD contained cdLC1 and were identified as cdLC1+actin, cdLC1+HC(s1), cdLC1+actin+HC(s1), and cdLC1+two actins+HC(s1), respectively. The same products were observed when cross-linking was performed in cardiac myofibrils incubated with cdS1. The production of cross-linked complexes of the heavy and light chain with actin decreased with an increase in the molar ratio of cdS1:actin. To test whether the orientation of myosin heads depended on a degree of occupation of thin filaments, myofibrils were irrigated with varying concentrations of cdS1. Fluorescence polarization measurements of cdS1 bound to individual t- bands revealed that the orientation depended on the concentration.
KW - Cardiac muscle
KW - Cross-linking
KW - F-actin
KW - Fluorescence
KW - Polarization
UR - http://www.scopus.com/inward/record.url?scp=0030683682&partnerID=8YFLogxK
U2 - 10.1161/01.RES.81.5.688
DO - 10.1161/01.RES.81.5.688
M3 - Article
C2 - 9351442
AN - SCOPUS:0030683682
SN - 0009-7330
VL - 81
SP - 688
EP - 693
JO - Circulation Research
JF - Circulation Research
IS - 5
ER -