title: Properties of the alkali light-chain-20-kilodalton fragment complex from skeletal myosin heads.
abstract: We have developed a rapid and reproducible procedure widely applicable to the preparation of pure aqueous solutions of the complex between an alkali light chain and the COOH-terminal heavy-chain fragments of skeletal myosin chymotryptic subfragment 1 (S-1) split by various proteases. It was founded on the remarkable ethanol solubility of these complexes. A systematic study of the ethanol fractionation of the tryptic (27K-50K-20K)-S-1 (A2) showed the NH2-terminal 27K fragment to behave like a specific protein entity being quantitatively precipitated at a relatively low ethanol concentration. Only the 20K peptide-A2 complex remained in solution when the S-1 derivative was treated with exactly 4 volumes of ethanol in the presence of 6 M guanidinium chloride. At a lower ethanol concentration, a soluble mixture of 50K and 20K peptides together with the light chain was obtained. The isolated 20K fragment-A2 system containing a 1:1 molar ratio of each component was investigated by biochemical and 1H nuclear magnetic resonance (NMR) techniques to highlight its structure and the interaction of the 20K heavy-chain segment with F-actin and with the light chain. During the treatment of the complex with alpha-chymotrypsin, only the 20K peptide was fragmented in contrast to its stability within the whole S-1. The binding of F-actin to the complex led, however, to a strong inhibition of its chymotryptic degradation. 1-Ethyl-3-[3-(dimethylamino)propyl]carbodiimide cross-linking of F-actin to the complex produced covalent actin-20K peptide only, the amount of which was lower relative to that observed with the entire split S-1.(ABSTRACT TRUNCATED AT 250 WORDS)
mesh_terms: D000199:Actins / Q000378:metabolism; D000818:Animals; D002918:Chymotrypsin / Q000378:metabolism; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy / Q000379:methods; D009132:Muscles / Q000378:metabolism; D015879:Myosin Subfragments; D009218:Myosins / Q000302:isolation & purification / Q000378:metabolism*; D010446:Peptide Fragments / Q000302:isolation & purification / Q000378:metabolism*; D011485:Protein Binding; D011817:Rabbits; D014357:Trypsin / Q000378:metabolism
chemical_list: D000199:Actins; D015879:Myosin Subfragments; D010446:Peptide Fragments; D002918:Chymotrypsin; D014357:Trypsin; D009218:Myosins