A Variable Domain near the ATP-Binding Site in Drosophila Muscle Myosin Is Part of the Communication Pathway between the Nucleotide and Actin-binding Sites

Miller, Becky M.; Bloemink, Marieke J.; Nyitrai, Miklós; Bernstein, Sanford I.; Geeves, Michael A.

doi:10.1016/j.jmb.2007.02.042

Cited by 26 publications

(31 citation statements)

References 45 publications

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“…Note that these observations mirror those from studies of the alternatively spliced exon 7 of Drosophila melanogaster myosin II (27). Those investigators demonstrated a similar influence of substitutions within the upper 50-kDa subdomain on nucleotide binding and release kinetics.…”

Section: Discussionsupporting

confidence: 77%

Role of Insert-1 of Myosin VI in Modulating Nucleotide Affinity

Pylypenko

Song

Squires

et al. 2011

Journal of Biological Chemistry

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Myosin VI is unique in its directionality among myosin superfamily members and also displays a slow and strain-dependent rate of ATP binding that allows for gating between its heads. In this study we demonstrate that leucine 310 is positioned by a class VI-specific insert, insert-1, so as to account for the selective hindrance of ATP versus ADP binding. Mutation of leucine 310 to glycine removes all influence of insert-1 on ATP binding. Furthermore, by analyzing myosin VI structures with either leucine 310 substituted to a glycine or complete removal of insert-1, we conclude that nucleotides may initially bind to myosin by their purine rings before docking their phosphate moieties. Otherwise, insert-1 could not exert a differential influence on ATP versus ADP binding.Myosin VI is the only myosin superfamily member that has been shown to traffic toward the minus (pointed) end of actin filaments (1). This unique directionality is made possible due to a myosin VI class-specific structural element that is known as insert-2 and enables myosin VI to perform unique cellular functions (2). This insert wraps around the myosin VI converter and binds a calmodulin via an unusual calmodulin binding motif (3). This redirects the lever arm of myosin VI ϳ120°as compared with plus-end-directed myosin motors.Like myosin V, dimeric myosin VI can function as a processive motor on actin (4 -7). That is to say that the dimer can move along actin in a hand-over-hand fashion with steps of 30 -36 nm. As in the case of the plus-end directed processive dimer of myosin Va (8 -11), the processive movement of myosin VI is optimized by "gating" between the heads. Intramolecular strain stalls the lead head until the rear head has detached from actin as the dimer moves along the filament (8 -12). However, the mechanism of how this is accomplished differs between myosin V and VI, necessitated by their opposite directionalities. In the case of myosin V, intramolecular strain greatly decreases the rate of dissociation of MgADP from the lead head (8 -11), whereas in the case of myosin VI, ADP dissociation is unaffected, but ATP binding is greatly slowed on the lead head, preventing its detachment (12). This is illustrated in Fig. 1.Based on the structure of myosin VI (3), it appeared that another myosin VI class-specific insert, insert-1, is responsible for impeding ATP binding. Indeed, even in the absence of strain, binding of ATP to monomeric myosin VI is at least an order of magnitude slower than binding to myosin V (3), and removal of the 26-residue insert increased the rate of ATP binding (3, 12). The mechanism of how this is achieved was not entirely clear, although we postulated that insert-1 positioned the side chain of leucine 310 physically in the way so that it impeded ATP but not ADP entry into the pocket.In this study we examine if positioning of leucine 310 does indeed provide steric hindrance for ATP binding and is in fact the mechanism by which myosin VI gating is achieved. Although we have examined the effects of insert-1 on nucleo...

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Section: Discussionsupporting

confidence: 77%

Role of Insert-1 of Myosin VI in Modulating Nucleotide Affinity

Pylypenko

Song

Squires

et al. 2011

Journal of Biological Chemistry

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“…Finally, we previously identified a region of the upper 50 kDa that is different between the α and β isoforms and could be important for the communication between the nucleotide and actin-binding sites (18). This region is labeled in the alignment as the exon 7 region by analogy with Drosophila muscle myosins II because this area is alternately spliced in the fly to generate different myosin II isoforms from a single gene (27). This region shows variation among not only the four fast isoforms but also the cardiac and EO isoforms.…”

Section: Resultsmentioning

confidence: 99%

“…The alignments were submitted to the alignment interface of SWISS-MODEL, and the generated models were validated using WHAT CHECK (26). A sequence identity of 60% (±0.5%) was found when aligning the human skeletal myosin head sequences with class II scallop myosin, allowing us to build well resolved homology models (27). The scallop myosin structures used as templates were chosen because they represent various conformational states of myosin during the cross-bridge cycle: the post-power stroke state containing ADP-BeF x in the nucleotide pocket (Protein Data Bank code 1kk8), the pre-power stroke state containing ADP-VO 4 (Protein Data Bank code 1qvi), and one scallop myosin structure without a nucleotide in the binding pocket (Protein Data Bank code 1sr6) representing the near rigor state of myosin.…”

Section: Methodsmentioning

confidence: 99%

The Superfast Human Extraocular Myosin Is Kinetically Distinct from the Fast Skeletal IIa, IIb, and IId Isoforms

Bloemink

Deacon

Resnicow

et al. 2013

Journal of Biological Chemistry

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Background: Characterization of individual muscle myosin isoforms has been limited by the availability of pure samples of isoforms.Results: Human myosin isoforms expressed in mouse cell line all have different ADP affinities.Conclusion: The superfast extraocular myosin is kinetically distinct from the fast skeletal IIa, IId, and IIb isoforms.Significance: This is the first time adult human sarcomeric myosins have been isolated and characterized by transient kinetics.

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“…This observation suggests that the u50 domain and switch 1 move simultaneously, consistent with the present observation that they belong to the same semirigid unit. Moreover, Geeves and coworkers (23) showed that it is difficult to weaken the switch-1 to u50 domain interaction by a few single point mutations, indicating that switch 1 is firmly embedded in the u50 domain.…”

Section: Discussionmentioning

confidence: 99%

Structural mechanism of the ATP-induced dissociation of rigor myosin from actin

Kühner

Fischer

2011

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

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Myosin is a true nanomachine, which produces mechanical force from ATP hydrolysis by cyclically interacting with actin filaments in a four-step cycle. The principle underlying each step is that structural changes in separate regions of the protein must be mechanically coupled. The step in which myosin dissociates from tightly bound actin (the rigor state) is triggered by the 30 Å distant binding of ATP. Large conformational differences between the crystal structures make it difficult to perceive the coupling mechanism. Energetically accessible transition pathways computed at atomic detail reveal a simple coupling mechanism for the reciprocal binding of ATP and actin.chemomechanical coupling | conformational transition | conjugate peak refinement | muscle contraction | power-stroke M otility as well as intracellular transport are essential functions in all organisms, from unicellulars to highest eukaryotes. They are driven by molecular motors, which convert the chemical energy of ATP hydrolysis into mechanical force (1). One family of motors are myosins, which drive processes like cytokinesis, vesicle transport, and muscle contraction by walking along actin fibrils (2).The N-terminal globular domain of myosin (called the head) contains all the functional domains (i.e., the ATP binding site, the actin-binding regions, and the rotating "converter" domain). It is able to hydrolyze ATP and move along an actin filament on its own (3). The cyclic interaction of myosin and actin to create motion is known as the Lymn-Taylor cycle (4) (Fig. 1A). The principle underlying each of the four Lymn-Taylor steps is that structural change in one region of the protein is tightly coupled to change in another domain of the protein. This sort of welldefined coupling between parts of the motor really is the essence of any type of motor (for instance, the motions of the valve and the piston are coupled in a car engine). The key to understanding how myosin works as a nanomachine is to understand in each Lymn-Taylor step how the structural information is communicated from one part of myosin to another (much like finding the crankshaft and the timing belt in the engine analogy).The focus here is on the rigor-dissociation step (Fig. 1A, I → II), in which, upon binding ATP, the head dissociates from the actin fibril and goes from the rigor state (thus called for the stiffness of corpses when ATP gets depleted in muscle cells) to the prerecovery (also known as postrigor) state (kinetic relaxation time approximately 10 ms in excess ATP; ref. 5). In this step, small changes in the ATP binding site are coupled to large structural changes in the 30-Å distant actin-binding domain that substantially reduce binding affinity for actin (6, 7), thus leading to the dissociation. We describe that mechanism at atomic detail using simulations of the conformational transition between the rigor and the postrigor states and present molecular movies of the energetically accessible transition pathways (SI Text). Fig. 1B shows the rigor-like crystal structure of...

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A Variable Domain near the ATP-Binding Site in Drosophila Muscle Myosin Is Part of the Communication Pathway between the Nucleotide and Actin-binding Sites

Cited by 26 publications

References 45 publications

Role of Insert-1 of Myosin VI in Modulating Nucleotide Affinity

Role of Insert-1 of Myosin VI in Modulating Nucleotide Affinity

The Superfast Human Extraocular Myosin Is Kinetically Distinct from the Fast Skeletal IIa, IIb, and IId Isoforms

Structural mechanism of the ATP-induced dissociation of rigor myosin from actin

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