Molecular movements in the actomyosin complex: F-actin-promoted internal cross-linking of the 25- and 20-kDa heavy chain fragments of skeletal myosin subfragment.
We describe, for the first time, the F-actin-promoted changes in the spatial relationship of strands in the NH2-terminal 25-kDa and COOH-terminal 20-kDa heavy chain fragments of the skeletal myosin subfragment 1 (S-1), detected by their exclusive chemical cross-linking in the rigor F-actin-S-1 complex with m-maleimidobenzoic acid N-hydroxysuccinimide ester (MBS). Quantitative electrophoretic analysis of the reaction products showed extensive conversion of the 95-kDa heavy chain of the actin-bound S-1 into a new species with an apparent mass of 135 kDa (yield = 50-60%), whereas the heavy chain mobility remained unaffected when actin was omitted. The 135-kDa entity retained the fluorescence of AEDANS-S-1 but not of AEDANS-actin, indicating that it was not a cross-linked acto-heavy chain adduct. Its extent of production depended markedly on the S-1: actin molar ratio and was maximum near a ratio of 1:4. The MBS treatment of acto-S-1 led also to some covalent actin-actin oligomers which could be suppressed by using trypsin-truncated F-actin lacking Cys-374, without altering the generation of the 135-kDa heavy chain derivative.(ABSTRACT TRUNCATED AT 250 WORDS)
Bertrand
R
R
Centre de Recherches de Biochimie Macromoléculaire du CNRS, INSERM, U 249 Université de Montpellier I, France.
Derancourt
J
J
Kassab
R
R
D000199:Actins / Q000737:chemistry*; D000205:Actomyosin / Q000737:chemistry*; D000818:Animals; D003432:Cross-Linking Reagents; D004591:Electrophoresis, Polyacrylamide Gel; D005456:Fluorescent Dyes; D009132:Muscles / Q000737:chemistry; D015879:Myosin Subfragments / Q000737:chemistry*; D009282:Naphthalenesulfonates / Q000737:chemistry; D011817:Rabbits; D013388:Succinimides / Q000737:chemistry; D013438:Sulfhydryl Compounds / Q000737:chemistry
D000199:Actins; D003432:Cross-Linking Reagents; D005456:Fluorescent Dyes; D015879:Myosin Subfragments; D009282:Naphthalenesulfonates; D013388:Succinimides; D013438:Sulfhydryl Compounds; C048876:1,5-AEDANS; C013097:3-maleimidobenzoyl N-hydroxysuccinimide; D000205:Actomyosin