Logo of pnasPNASInfo for AuthorsSubscriptionsAboutThis Article
Proc Natl Acad Sci U S A. 1996 Mar 19; 93(6): 2285–2289.

The role of surface loops (residues 204-216 and 627-646) in the motor function of the myosin head.


A characteristic feature of all myosins is the presence of two sequences which despite considerable variations in length and composition can be aligned with loops 1 (residues 204-216) and 2 (residues 627-646) in the chicken myosin-head heavy chain sequence. Recently, an intriguing hypothesis has been put forth suggesting that diverse performances of myosin motors are achieved through variations in the sequences of loops 1 and 2 [Spudich, J. (1994) Nature (London) 372, 515-518]. Here, we report on the study of the effects of tryptic digestion of these loops on the motor and enzymatic functions of myosin. Tryptic digestions of myosin, which produced heavy meromyosin (HMM) with different percentages of molecules cleaved at both loop 1 and loop 2, resulted in the consistent decrease in the sliding velocity of actin filaments over HMM in the in vitro motility assays, did not affect the Vmax, and increased the Km values for actin-activated ATPase of HMM. Selective cleavage of loop 2 on HMM decreased its affinity for actin but did not change the sliding velocity of actin in the in vitro motility assays. The cleavage of loop 1 and HMM decreased the mean sliding velocity of actin in such assays by almost 50% but did not alter its affinity for HMM. To test for a possible kinetic determinant of the change in motility, 1-N6-ethenoadenosine diphosphate (epsilon-ADP) release from cleaved and uncleaved myosin subfragment 1 (S1) was examined. Tryptic digestion of loop 1 slightly accelerated the release of epsilon-ADP from S1 but did not affect the rate of epsilon-ADP release from acto-S1 complex. Overall, the results of this work support the hypothesis that loop 1 can modulate the motor function of myosin and suggest that such modulation involves a mechanism other than regulation of ADP release from myosin.

Full text

Full text is available as a scanned copy of the original print version. Get a printable copy (PDF file) of the complete article (1.2M), or click on a page image below to browse page by page. Links to PubMed are also available for Selected References.

Images in this article

Click on the image to see a larger version.

Selected References

These references are in PubMed. This may not be the complete list of references from this article.
  • Ma YZ, Taylor EW. Kinetic mechanism of myofibril ATPase. Biophys J. 1994 May;66(5):1542–1553. [PMC free article] [PubMed]
  • Rayment I, Rypniewski WR, Schmidt-Bäse K, Smith R, Tomchick DR, Benning MM, Winkelmann DA, Wesenberg G, Holden HM. Three-dimensional structure of myosin subfragment-1: a molecular motor. Science. 1993 Jul 2;261(5117):50–58. [PubMed]
  • Sutoh K. Mapping of actin-binding sites on the heavy chain of myosin subfragment 1. Biochemistry. 1983 Mar 29;22(7):1579–1585. [PubMed]
  • Chaussepied P, Morales MF. Modifying preselected sites on proteins: the stretch of residues 633-642 of the myosin heavy chain is part of the actin-binding site. Proc Natl Acad Sci U S A. 1988 Oct;85(20):7471–7475. [PMC free article] [PubMed]
  • Yamamoto K. Shift of binding site at the interface between actin and myosin. Biochemistry. 1990 Jan 23;29(3):844–848. [PubMed]
  • Cheung P, Reisler E. Synthetic peptide of the sequence 632-642 on myosin subfragment 1 inhibits actomyosin ATPase activity. Biochem Biophys Res Commun. 1992 Dec 15;189(2):1143–1149. [PubMed]
  • Warrick HM, Spudich JA. Myosin structure and function in cell motility. Annu Rev Cell Biol. 1987;3:379–421. [PubMed]
  • Spudich JA. How molecular motors work. Nature. 1994 Dec 8;372(6506):515–518. [PubMed]
  • Uyeda TQ, Ruppel KM, Spudich JA. Enzymatic activities correlate with chimaeric substitutions at the actin-binding face of myosin. Nature. 1994 Apr 7;368(6471):567–569. [PubMed]
  • Umemoto S, Sellers JR. Characterization of in vitro motility assays using smooth muscle and cytoplasmic myosins. J Biol Chem. 1990 Sep 5;265(25):14864–14869. [PubMed]
  • Shimizu T, Furusawa K, Ohashi S, Toyoshima YY, Okuno M, Malik F, Vale RD. Nucleotide specificity of the enzymatic and motile activities of dynein, kinesin, and heavy meromyosin. J Cell Biol. 1991 Mar;112(6):1189–1197. [PMC free article] [PubMed]
  • Mornet D, Pantel P, Audemard E, Kassab R. The limited tryptic cleavage of chymotryptic S-1: an approach to the characterization of the actin site in myosin heads. Biochem Biophys Res Commun. 1979 Aug 13;89(3):925–932. [PubMed]
  • Mornet D, Bertrand RU, Pantel P, Audemard E, Kassab R. Proteolytic approach to structure and function of actin recognition site in myosin heads. Biochemistry. 1981 Apr 14;20(8):2110–2120. [PubMed]
  • Bálint M, Wolf I, Tarcsafalvi A, Gergely J, Sréter FA. Location of SH-1 and SH-2 in the heavy chain segment of heavy meromyosin. Arch Biochem Biophys. 1978 Oct;190(2):793–799. [PubMed]
  • Godfrey JE, Harrington WF. Self-association in the myosin system at high ionic strength. I. Sensitivity of the interaction to pH and ionic environment. Biochemistry. 1970 Feb 17;9(4):886–893. [PubMed]
  • Spudich JA, Watt S. The regulation of rabbit skeletal muscle contraction. I. Biochemical studies of the interaction of the tropomyosin-troponin complex with actin and the proteolytic fragments of myosin. J Biol Chem. 1971 Aug 10;246(15):4866–4871. [PubMed]
  • Weeds AG, Pope B. Studies on the chymotryptic digestion of myosin. Effects of divalent cations on proteolytic susceptibility. J Mol Biol. 1977 Apr;111(2):129–157. [PubMed]
  • Margossian SS, Lowey S. Preparation of myosin and its subfragments from rabbit skeletal muscle. Methods Enzymol. 1982;85(Pt B):55–71. [PubMed]
  • Mocz G, Szilagyi L, Chen Lu R, Fabian F, Balint M, Gergely J. Effect of nucleotides, divalent cations and temperature on the tryptic susceptibility of myosin subfragment 1. Eur J Biochem. 1984 Dec 3;145(2):221–229. [PubMed]
  • Laemmli UK. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. [PubMed]
  • Homsher E, Wang F, Sellers JR. Factors affecting movement of F-actin filaments propelled by skeletal muscle heavy meromyosin. Am J Physiol. 1992 Mar;262(3 Pt 1):C714–C723. [PubMed]
  • Rosenfeld SS, Taylor EW. The dissociation of 1-N6-ethenoadenosine diphosphate from regulated actomyosin subfragment 1. J Biol Chem. 1987 Jul 25;262(21):9994–9999. [PubMed]
  • Phan BC, Faller LD, Reisler E. Kinetic and equilibrium analysis of the interactions of actomyosin subfragment-1.ADP with beryllium fluoride. Biochemistry. 1993 Aug 3;32(30):7712–7719. [PubMed]
  • Taylor EW. Transient phase of adenosine triphosphate hydrolysis by myosin, heavy meromyosin, and subfragment 1. Biochemistry. 1977 Feb 22;16(4):732–739. [PubMed]
  • Johnson KA, Taylor EW. Intermediate states of subfragment 1 and actosubfragment 1 ATPase: reevaluation of the mechanism. Biochemistry. 1978 Aug 22;17(17):3432–3442. [PubMed]
  • Ueno H, Harrington WF. An enzyme-probe study of motile domains in the subfragment-2 region of myosin. J Mol Biol. 1984 Dec 15;180(3):667–701. [PubMed]
  • Maita T, Hayashida M, Tanioka Y, Komine Y, Matsuda G. The primary structure of the myosin head. Proc Natl Acad Sci U S A. 1987 Jan;84(2):416–420. [PMC free article] [PubMed]
  • Yamamoto K. Identification of the site important for the actin-activated MgATPase activity of myosin subfragment-1. J Mol Biol. 1991 Jan 20;217(2):229–233. [PubMed]
  • Hynes TR, Block SM, White BT, Spudich JA. Movement of myosin fragments in vitro: domains involved in force production. Cell. 1987 Mar 27;48(6):953–963. [PubMed]
  • Lowey S, Waller GS, Trybus KM. Function of skeletal muscle myosin heavy and light chain isoforms by an in vitro motility assay. J Biol Chem. 1993 Sep 25;268(27):20414–20418. [PubMed]
  • Siemankowski RF, Wiseman MO, White HD. ADP dissociation from actomyosin subfragment 1 is sufficiently slow to limit the unloaded shortening velocity in vertebrate muscle. Proc Natl Acad Sci U S A. 1985 Feb;82(3):658–662. [PMC free article] [PubMed]
  • Marston SB, Taylor EW. Comparison of the myosin and actomyosin ATPase mechanisms of the four types of vertebrate muscles. J Mol Biol. 1980 Jun 5;139(4):573–600. [PubMed]
  • Taylor EW. Kinetic studies on the association and dissociation of myosin subfragment 1 and actin. J Biol Chem. 1991 Jan 5;266(1):294–302. [PubMed]
  • Ohichi T, Hozumi T, Higashi-Fujime S. In vitro motility of proteolytically cleaved myosin subfragment 1 on a lysine-coated surface. J Biochem. 1993 Sep;114(3):299–302. [PubMed]
  • Botts J, Muhlrad A, Takashi R, Morales MF. Effects of tryptic digestion on myosin subfragment 1 and its actin-activated adenosinetriphosphatase. Biochemistry. 1982 Dec 21;21(26):6903–6905. [PubMed]
  • Rosenfeld SS, Taylor EW. Reactions of 1-N6-ethenoadenosine nucleotides with myosin subfragment 1 and acto-subfragment 1 of skeletal and smooth muscle. J Biol Chem. 1984 Oct 10;259(19):11920–11929. [PubMed]
  • White HD, Belknap B, Jiang W. Kinetics of binding and hydrolysis of a series of nucleoside triphosphates by actomyosin-S1. Relationship between solution rate constants and properties of muscle fibers. J Biol Chem. 1993 May 15;268(14):10039–10045. [PubMed]

Articles from Proceedings of the National Academy of Sciences of the United States of America are provided here courtesy of National Academy of Sciences


Related citations in PubMed

See reviews...See all...

Cited by other articles in PMC

See all...


  • PubMed
    PubMed citations for these articles
  • Substance
    PubChem Substance links

Recent Activity

Your browsing activity is empty.

Activity recording is turned off.

Turn recording back on

See more...