Amino Acid Sequence of the 203-Residue Fragment of the Heavy Chain of Chicken Gizzard Myosin Containing the SH Cysteine Residue12

A fluorescent fragment of Mr=23,800 was obtained by the papain digestion of N-iodoacetyl-N′-(5-sulfo-1-naphthyl)ethylene diamine (abbreviated as IAEDANS)-modified chicken gizzard myosin. The fragment was isolated by gel filtration on a Sephadex G-100 column in the presence of 5 M guanidine-HCI followed by anion exchange chromatography on a QAE Sephadex A-50 column. This fragment contained 203 amino acid residues which could be assigned as a COOH-terminal part of the S-1 heavy chain based on the homology with the known sequence of rabbit skeletal myosin fragment. The amino acid sequence was K-G-M-F-R-T-V-G-Q-L-Y-K-E-Q-L-T-K-L-M-T20-T-L-R-N-T-N-P-N-F-V-R-C-I-I-P-N-H-E-K-R40-A-G-K-L-D-A-H-L-V-L-E-Q-L-R-C-N-G-V-L-E60-G-I-R-I-C-R-Q-G-F-P-N-R-I-V-F-Q-E-F-R-Q80-R-Y-E-I-L-A-A-N-A-I-P-K-G-F-M-D-G-K-Q-A100-C-I-L-M-I-K-A-L-E-L-D-P-N-L-Y-R-I-G-Q-S120-K-J-F-F-R-T-G-V-L-A-H-L-E-E-E-R-D-L-K-I140-T-D-V-I-I-A-F-Q-A-Q-C-R-G-Y-L-A-R-K-A-F160-A-K-R-Q-Q-Q-L-T-A-M-K-V-I-Q-R-N-C-A-A-Y180-L-K-L-R-N-W-Q-W-W-R-L-F-T-K-V-K-P-L-L-Q200-V-T-R⁠. The cysteine residue which was modified with IAEDANS was of the SH1 type (Cys-65). Pro-197 was suggested to be the NHrterminal boundary of the alpha-helical coiled-coil rod sequence of gizzard myosin, based on the homology with the nematode sequence reported by MacLachlan and Karn (Proc. Natl. Acad. Sd. U.S. 80, 4253–4257 (1983)). Three different COOH-terminal peptides (Val-Lys-Pro-Leu-Leu-Gln-Val Thr-Arg, Val-Lys-Pro-Leu-Leu-Gln, and Val-Lys-Pro-Leu-Leu) were isolated from the tryptic digest of this fragment, suggesting that three close peptide bonds in the myosin neck (Leu 198-Gln 199, Gln 199-Val 200, and Arg 203-) are susceptible to papain digestion. The COOH-terminal 53-residue sequence of this fragment, constituting the neck part of myosin, contained 13 positively charged residues but no negatively charged residue. The possible role of the positive charges in the binding of the heavy chain to the regulatory light chain is discussed.