Location of SH-1 and SH-2 in the heavy chain segment of heavy meromyosin

Bálint, M.; Wolf, Imre; Tarcsafalvi, Adel; Gergely, J.; Sreter, F. A.

doi:10.1016/0003-9861(78)90339-9

Cited by 192 publications

(114 citation statements)

References 18 publications

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“…We showed previously that the tryptic digestion of skeletal S1 and ␤S1 splits the heavy chains at two points, Loops 1 and 2, producing the characteristic 25-, 50-, and 20-kDa fragments (32,52) (Fig. 1).…”

Section: Tryptic Digestion Of the C-loop In ␤S1-mentioning

confidence: 83%

The Myosin Cardiac Loop Participates Functionally in the Actomyosin Interaction

Ajtai

Garamszegi

Watanabe

et al. 2004

Journal of Biological Chemistry

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The motor protein myosin in association with actin transduces chemical free energy in ATP into work in the form of actin translation against an opposing force. Mediating the actomyosin interaction in myosin is an actin binding site distributed among several peptides on the myosin surface including surface loops contributing to affinity and actin regulation of myosin ATPase. A structured surface loop on ␤-cardiac myosin, the cardiac or C-loop, was recently demonstrated to affect myosin ATPase and was indirectly implicated in the actomyosin interaction. The C-loop is a conserved feature of all myosin isoforms with crystal structures, suggesting that it is an essential part of the core energy transduction machinery. It is shown here that proteolytic digestion of the C-loop in ␤-cardiac myosin eliminates actin-activated myosin ATPase and reduces actomyosin affinity in rigor more than 100-fold. Studies of C-loop function in smooth muscle myosin were also undertaken using sitedirected mutagenesis. Mutagenesis of a single charged residue in the C-loop of smooth muscle myosin alters actomyosin affinity and doubles myosin in vitro motility and actin-activated ATPase velocities, thereby involving a charged region of the loop in the actomyosin interaction. It appears likely that the C-loop is an essential electrostatic binding site for actin involved in modulation of actomyosin affinity and regulation of actomyosin ATPase velocity.The motor protein myosin is an ATPase and an actin-binding protein transducing ATP free energy into work. In muscle, myosin, actin, and ATP constitute the unitary work producer. The cyclical interaction of these components, a sequence of states each characterized as a static relation between the proteins and an intermediate in the degradation of ATP, is a contraction cycle. In a contraction cycle, myosin hydrolyzes ATP in its active site and forms a weak association with actin at a separate actin binding site. Release of phosphate from the active site initiates strong binding to actin and a large conformation change in myosin that produces work in the form of actin translation against a load. Several solved crystal structures represent intermediates in ATP hydrolysis and define myosin conformation changes in the absence of actin (1-6). Presently, we lack a detailed understanding of actomyosin structure and in particular how the elements making up the actomyosin interface contribute to mutual affinity, actin regulation of phosphate release, and the ability of myosin-bound nucleotide to modulate actomyosin affinity.Myosin consists of a globular head domain containing the enzymatic portion of the molecule and a tail involved in filament assembly. The globular head separated from its tail portion is called subfragment 1 (S1). 1 S1 interacts with filamentous actin (F-actin) consisting of polymerized actin monomers. An atomic model for F-actin built from monomeric actin crystal structures (7-9) satisfies x-ray diffraction data restraints (10) and is consistent with electron microscopic imagery (11). In...

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Section: Tryptic Digestion Of the C-loop In ␤S1-mentioning

confidence: 83%

The Myosin Cardiac Loop Participates Functionally in the Actomyosin Interaction

Ajtai

Garamszegi

Watanabe

et al. 2004

Journal of Biological Chemistry

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“…1) (3). Two regions corresponding to the 25/50-kDa and 50/20-kDa junctions were not resolved in the crystal structure of chicken skeletal myosin S1, suggesting that they might exist as flexible surface loops (4).…”

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confidence: 99%

Functional Characterization of Vertebrate Nonmuscle Myosin IIB Isoforms Using Dictyostelium Chimeric Myosin II

Takahashi

Hiratsuka

et al. 2001

Journal of Biological Chemistry

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The alternatively spliced isoform of nonmuscle myosin II heavy chain B (MHC-IIB) with an insert of 21 amino acids in the actin-binding surface loop (loop 2), MHC-IIB(B2), is expressed specifically in the central nervous system of vertebrates. To examine the role of the B2 insert in the motor activity of the myosin II molecule, we expressed chimeric myosin heavy chain molecules using the Dictyostelium myosin II heavy chain as the backbone. We replaced the Dictyostelium native loop 2 with either the noninserted form of loop 2 from human MHC-IIB or the B2-inserted form of loop 2 from human MHC-IIB(B2). The transformant Dictyostelium cells expressing only the B2-inserted chimeric myosin formed unusual fruiting bodies. We then assessed the function of chimeric proteins, using an in vitro motility assay and by measuring ATPase activities and binding to F-actin. We demonstrate that the insertion of the B2 sequence reduces the motor activity of Dictyostelium myosin II, with reduction of the maximal actin-activated ATPase activity and a decrease in the affinity for actin. In addition, we demonstrate that the native loop 2 sequence of Dictyostelium myosin II is required for the regulation of the actin-activated ATPase activity by phosphorylation of the regulatory light chain.

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“…We have previously shown that tryptic digestion at 0°C of LC2-deficient papain S1 generates the C-terminal heavy chain fragment of 26 kDa instead of a 20 kDa fragment formed at room temperature [6]. The latter seems to correspond to the C-terminal 20 kDa tryptic fragment of the heavy chain of chymotryptic SI routinely prepared by digestion of myosin at room temperature [10]. As shown in fig.2, tryptic cleavage of SI prepared by digestion of myosin at 0°C, in addition to the N-terminal 27 kDa and central 50 kDa fragments, generated two fluorescent C-terminal fragments, of 20 kDa and 23 kDa, the latter being the predominant species.…”

Section: Resultsmentioning

confidence: 99%

Dependence of the length of the heavy chain of chymotryptic subfragment 1 on the temperature of myosin digestion

Pliszka

Rędowicz

Strzelecka‐Gołaszewska

1989

FEBS Letters

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Limited digestion of filamentous myosin with chymotrypsin at 0°C in the absence of divalent cations generates two forms of subfragment 1 (SI), with heavy chains of 95 kDa and 98 kDa. The difference is at the C-terminal end of the chain. The 98 kDa form prevails, in contrast to the preparations obtained by digestion at room temperature which consist of the shorter species and only traces of the longer one. The results support the idea of a temperature-dependent conformational transition at the head-rod junctional region of the myosin heavy chain.

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Location of SH-1 and SH-2 in the heavy chain segment of heavy meromyosin

Cited by 192 publications

References 18 publications

The Myosin Cardiac Loop Participates Functionally in the Actomyosin Interaction

The Myosin Cardiac Loop Participates Functionally in the Actomyosin Interaction

Functional Characterization of Vertebrate Nonmuscle Myosin IIB Isoforms Using Dictyostelium Chimeric Myosin II

Dependence of the length of the heavy chain of chymotryptic subfragment 1 on the temperature of myosin digestion

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