Antibodies directed against N-terminal residues on actin do not block acto-myosin binding
Several studies using a variety of approaches have suggested a possible role for the amino-terminal residues of skeletal muscle actin in acto-myosin interaction. In order to assess the significance of acto-S-1 contacts involving the N-terminal segment of actin, the authors have prepared polyclonal antisera against a synthetic /sup 14/C-peptide corresponding to the seven amino-terminal residues of rabbit skeletal muscle actin (..cap alpha..-N-terminal peptide). Affinity-purified immunoglobulin (Ig) G (and Fab) prepared from these antisera reacts strongly and specifically with the amino-terminal segment of both G- and F-actin but not with myosin subfragment 1 (S-1). This specificity was determined by Western blot analysis of actin and its proteolytic fragments and the inhibition of the above reactivity by the ..cap alpha..-N-terminal peptide. The ..cap alpha..-N-terminal peptide did not interact with S-1 in solution, affect S-1 and actin-activated S-1 MgATPase, or cause dissociation of the acto-S-1 complex. In separate experiments F-actin could be cosedimented with S-1 and affinity-purified IgG or Fab by using an air-driven ultracentrifuge. Densitometric analysis of sodium dodecyl sulfate/polyacrylamide gels of pellet and supernatant fractions from such experiments demonstrated the binding of both S-1 and IgG or Fab to the same F-actin protomer. The results suggest that, while the acidic N-terminal amino acids of actin may contact the myosin head, these residues cannot be the main determinants of acto-S-1 interaction.
- Research Organization:
- Univ. of California, Los Angeles
- OSTI ID:
- 5515208
- Journal Information:
- Biochemistry; (United States), Vol. 26:19
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
ACTIN
ENZYME IMMUNOASSAY
MOLECULAR STRUCTURE
CARBON 14 COMPOUNDS
ANTIBODIES
ATP-ASE
ELECTROPHORESIS
IMMUNOLOGY
MYOSIN
RABBITS
ACID ANHYDRASES
ANIMALS
ENZYMES
GLOBULINS
HYDROLASES
IMMUNOASSAY
LABELLED COMPOUNDS
MAMMALS
ORGANIC COMPOUNDS
PHOSPHOHYDROLASES
PROTEINS
VERTEBRATES
550201* - Biochemistry- Tracer Techniques