Identification of myosin-binding sites on the actin sequence

Sutoh, Kazuo

doi:10.1021/bi00258a020

Cited by 343 publications

(342 citation statements)

References 30 publications

(54 reference statements)

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“…In the present investigation, we have extended our previous study of the interaction between troponin-I and actin and, using both native F-actin and defined peptides of actin, have employed 'H-NMR spectroscopic techniques to identify the residues of rabbit skeletal muscle actin involved in the interaction with the inhibitory segment of troponin-I. Residues at the N-terminal region of actin are here shown to form a surface of contact on actin for troponin-I at a site on actin sequentially close to that previously found for the heavy chain of the skeletal myosin head [15,161. These data are discussed in relation to a structural mechanism for the regulation of actomyosin ATPase activity.…”

supporting

confidence: 55%

“…Several lines of experimental evidence have implicated the N-terminal residues of actin as providing a site of contact between actin and the heavy chain of myosin S1. Cross-linking studies [15,261 have shown that the Nterminal 12 residues of actin contain a site for EDC crosslinking to the 20-kDa and the 50-kDa fragments of myosin S1. The 20-kDa fragment has been shown to be the preferential site of cross-linked attachment to actin under a variety of conditions [27].…”

Section: Discussionmentioning

confidence: 99%

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The interaction of troponin‐I with the N‐terminal region of actin

Levine

Moir

Perry

1988

European Journal of Biochemistry

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The interaction between troponin-I and actin that underlies thin-filament regulation in striated muscle has been studied using proton magnetic resonance spectroscopy. A restricted portion of skeletal muscle troponin-I (residues 96-116) has previously been shown to be capable of inhibiting the MgATPase activity of actomyosin in a manner enhanced by tropomyosin [Syska et al. (1976) Biochem. J. 153,375 -3871. On the basis of homologous spectral effects for signals of specific groups observed in different complexes formed using the native proteins and a variety of defined peptides, it is concluded that the segment of troponin-I which has inhibitory activity interacts with the N-terminal region of actin. The surface of contact of the inhibitory segment of troponin-I with actin involves two regions of the N-terminal of actin. These are located between residues 1-7 and 19-44. The data are discussed in the context of a structural mechanism for the inhibition of myosin ATPase activation.Contraction of vertebrate skeletal muscle is regulated by the interaction of calcium with the troponin complex located on the actin-containing thin filaments [l -31. The binding of calcium to troponin-C releases the inhibition of the MgATPase activity of actomyosin by troponin-I that is bound to actin. Present evidence suggests that activation may occur by release of the inhibition of one of the kinetic steps of ATP hydrolysis rather than by merely reversing a steric impediment to myosin-actin interaction [3 -51. A series of structural changes transmitted through troponin-I and troponin-T, the tropomyosin-binding subunit, is implicit in triggering contraction. To understand more fully the mechanism of action of the troponin complex, precise information is required about the surfaces of contact between its subunits and the other thin filament proteins.

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supporting

confidence: 55%

Section: Discussionmentioning

confidence: 99%

The interaction of troponin‐I with the N‐terminal region of actin

Levine

Moir

Perry

1988

European Journal of Biochemistry

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“…Loops 2 and 3 are part of actin binding sites (9,10). Cross-linking studies suggested that positively charged residues in loops 2 and 3 interact with negatively charged residues in subdomain 1 of actin at the initial weakly bound state (11)(12)(13)(14)(15)(16)(17)(18)(19)(20)(21).…”

mentioning

confidence: 99%

Unique charge distribution in surface loops confers high velocity on the fast motor protein Chara myosin

Ito

Yamaguchi

Yanase

et al. 2009

Proc. Natl. Acad. Sci. U.S.A.

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Most myosins have a positively charged loop 2 with a cluster of lysine residues that bind to the negatively charged N-terminal segment of actin. However, the net charge of loop 2 of very fast Chara myosin is zero and there is no lysine cluster in it. In contrast, Chara myosin has a highly positively charged loop 3. To elucidate the role of these unique surface loops of Chara myosin in its high velocity and high actin-activated ATPase activity, we have undertaken mutational analysis using recombinant Chara myosin motor domain. It was found that net positive charge in loop 3 affected Vmax and Kapp of actin activated ATPase activity, while it affected the velocity only slightly. The net positive charge in loop 2 affected Kapp and the velocity, although it did not affect Vmax. Our results suggested that Chara myosin has evolved to have highly positively charged loop 3 for its high ATPase activity and have less positively charged loop 2 for its high velocity. Since high positive charge in loop 3 and low positive charge in loop 2 seem to be one of the reasons for Chara myosin's high velocity, we manipulated charge contents in loops 2 and 3 of Dictyostelium myosin (class II). Removing positive charge from loop 2 and adding positive charge to loop 3 of Dictyostelium myosin made its velocity higher than that of the wild type, suggesting that the charge strategy in loops 2 and 3 is widely applicable.actin ͉ ATPase ͉ motility ͉ cytoplasmic streaming ͉ molecular engineering C ytoplasmic streaming in characean algal cells is extremely fast, and this streaming is brought about by the movement of myosin-coated organelles along actin filament bundles fixed inside the cell (1). Myosin purified from Chara corallina could translocate actin filaments in the in vitro motility assay at a velocity comparable to that of the cytoplasmic streaming (approximately 50 m s Ϫ1 ) (2, 3). This velocity is about 10 times faster than that of the fast skeletal muscle myosin and the Chara myosin is the fastest motor protein known so far. We have cloned cDNA of the Chara myosin heavy chain (4) and succeeded in expressing functional motor domain (5, 6). The velocity of the expressed motor domain measured by in vitro motility assay was comparable to that of the native Chara myosin if we consider the difference in the lever arm length. Relevant steps in actomyosin ATPase cycle to the sliding velocity are ADP release from actomyosin and ATP reassociation resulting in actin-myosin dissociation. Kinetic analyses of Chara myosin motor domain revealed that both the ADP release and the ATP-induced dissociation from actin were very fast (6). Time spent in the strongly bound state with actin, which was estimated from these rates, was less than 1 ms. This very short strongly bound state, together with a large step size (7), are probably the reason for the very high velocity of Chara myosin. Actin-activated ATPase activity of expressed Chara motor domain was approximately 500 s Ϫ1 head Ϫ1

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“…In human skeletal muscle, the A1-type fast (ELC1f) and slow (ELC1s) ELC isoforms contain 40 -45 additional amino acids compared with the A2-type fast isoform (ELC3f) (14). The longer skeletal muscle ELC isoforms cross-link to the C-terminal region of actin through their N-terminal ␣-amino group and four lysines within the first 10 residues (10,(15)(16)(17). In human cardiac muscle, both ventricular (ELC1v) and atrial (ELC1a) ELC isoforms have N-terminal extensions of approximately the same length as their A1-type counterparts in skeletal muscle.…”

mentioning

confidence: 99%

The Essential Light Chain N-terminal Extension Alters Force and Fiber Kinetics in Mouse Cardiac Muscle

Miller

Palmer

Ruch

et al. 2005

Journal of Biological Chemistry

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The functional significance of the actin-binding region at the N terminus of the cardiac myosin essential light chain (ELC) remains elusive. In a previous experiment, the endogenous ventricular ELC was replaced with a protein containing a 10-amino acid deletion at positions 5-14 (ELC1v⌬5-14, referred to as 1v⌬5-14), a region that interacts with actin (Sanbe, A., Gulick, J., Fewell, J., and Robbins, J. (2001) J. Biol. Chem. 276, 32682-32686). 1v⌬5-14 mice showed no discernable mutant phenotype in skinned ventricular strips. However, because the myofilament lattice swells upon skinning, the mutant phenotype may have been concealed by the inability of the ELC to reach the actin-binding site. Using the same mouse model, we repeated earlier measurements and performed additional experiments on skinned strips osmotically compressed to the intact lattice spacing as determined by x-ray diffraction. 1v⌬5-14 mice exhibited decreased maximum isometric tension without a change in calcium sensitivity. The decreased force was most evident in 5-6-month-old mice compared with 13-15-month-old mice and may account for the greater ventricular wall thickness in young 1v⌬5-14 mice compared with age-matched controls. No differences were observed in unloaded shortening velocity at maximum calcium activation. However, 1v⌬5-14 mice exhibited a significant difference in the frequency at which minimum complex modulus amplitude occurred, indicating a change in cross-bridge kinetics. We hypothesize that the ELC N-terminal extension interaction with actin inhibits the reversal of the power stroke, thereby increasing isometric force. Our results strongly suggest that an interaction between residues 5-14 of the ELC N terminus and the C-terminal residues of actin enhances cardiac performance.

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Identification of myosin-binding sites on the actin sequence

Cited by 343 publications

References 30 publications

The interaction of troponin‐I with the N‐terminal region of actin

The interaction of troponin‐I with the N‐terminal region of actin

Unique charge distribution in surface loops confers high velocity on the fast motor protein Chara myosin

The Essential Light Chain N-terminal Extension Alters Force and Fiber Kinetics in Mouse Cardiac Muscle

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