what immediately follows the binding of atp to the myosin head

. 2020 Nov;598(22):5165-5182.

doi: 10.1113/JP280402. Epub 2020 Sep 9.

Myosin dynamics during relaxation in mouse soleus musculus and modulation past 2'-deoxy-ATP

Affiliations

• PMID: 32818298
• PMCID: PMC7719615
• DOI: 10.1113/JP280402

Gratis PMC article

Myosin dynamics during relaxation in mouse soleus muscle and modulation by 2'-deoxy-ATP

Weikang Ma  et al. J Physiol. 2020 Nov .

Free PMC commodity

Abstruse

Key points: Skeletal muscle relaxation has been primarily studied by assessing the kinetics of strength decay. Little is known about the resultant dynamics of structural changes in myosin heads during relaxation. The naturally occurring nucleotide 2-deoxy-ATP (dATP) is a myosin activator that enhances cross-bridge binding and kinetics. X-ray diffraction data indicate that with elevated dATP, myosin heads were extended closer to actin in relaxed muscle and myosin heads return to an ordered, resting country after wrinkle more apace. Molecular dynamics simulations of post-powerstroke myosin suggest that dATP induces structural changes in myosin heads that increase the expanse of the actin-bounden regions promoting myosin interaction with actin, which could explain the observed delays in the onset of relaxation. This study of the dATP-induced changes in myosin may be instructive for determining the structural changes desired for other potential myosin-targeted molecular compounds to treat musculus diseases.

Abstruse: Here nosotros used fourth dimension-resolved small-bending X-ray diffraction coupled with force measurements to report the structural changes in FVB mouse skeletal musculus sarcomeres during relaxation after tetanus contraction. To estimate the charge per unit of myosin deactivation, we followed the rate of the intensity recovery of the beginning-order myosin layer line (MLL1) and restoration of the resting spacing of the third and sixth social club of meridional reflection (Due south_M3 and Southward_M6 ) post-obit tetanic contraction. A transgenic mouse model with elevated skeletal muscle ii-deoxy-ATP (dATP) was used to written report how myosin activators may impact soleus muscle relaxation. X-ray diffraction bear witness indicates that with elevated dATP, myosin heads were extended closer to actin in resting musculus. Following contraction, there is a slight only significant filibuster in the decay of force relative to WT musculus while the render of myosin heads to an ordered resting land was initially slower, then became more rapid than in WT musculus. Molecular dynamics simulations of mail-powerstroke myosin suggest that dATP induces structural changes in myosin that increase the surface surface area of the actin-binding regions, promoting myosin interaction with actin. With dATP, myosin heads may remain in an activated state nigh the thin filaments following relaxation, accounting for the delay in force disuse and the initial delay in recovery of resting head configuration, and this could facilitate subsequent contractions.

Keywords: X-ray diffraction; dATP; molecular dynamics simulation; myosin; relaxation; skeletal muscle.

© 2020 The Authors. The Periodical of Physiology © 2020 The Physiological Society.

Conflict of interest statement

Competing interests

The authors declare no competing interests

Figures

Figure one.

A. Ten-ray diffraction patterns from relaxing (left panel) and contracting (correct panel) muscle at ten ms exposure. Fourth dimension class of equatorial intensity ratio (B) and lattice spacing (C) throughout activation and relaxation.

Figure 2.

A. Spacing of M3 reflection (S_M3); B. Radial width of M3 reflection (W_M3); C. Intensity of M3 reflection (I_M3); D. Spacing of M6 reflection (South_M6); E. Intensity of MLL1 reflection (I_MLL1) as a function of time during relaxation. Error bars are standard errors of the mean.

Figure 3.. Soleus musculus dATP content and the kinetics of forcefulness development in WT (black) and RNR (blue) soleus muscle.

A. Soleus muscle dATP content in WT and RNR soleus muscle (n = 11, unpaired T examination). B. The half time (Kd) and hill coefficient (h value) when the force arising profile fitted to colina function (n = 21 in WT and n = 24 in RNR, unpaired T test). C. MLL1 intensity changes at the beginning of electrical activation is not significantly different between WT and RNR mouse soleus muscle. D. Equatorial intensity ratio (I₁₁/I₁₀) as a part of average tetanic tension from WT and RNR mouse soleus muscle can be fitted into ane curve. In that location was no significant divergence in boilerplate tetanic tension between WT and RNR soleus muscle (inset, n = 22 in WT and n = 28 in RNR, unpaired T test).

Figure iv.. Normalized force and equatorial intensity ratio during tetanic contraction in WT (blackness) and RNR (blueish) muscle.

A. Time course of forcefulness during activation and relaxation from WT and RNR muscle. B. Relaxation kinetics from forcefulness decay. The time for the force drop to ninety% of the maximum tetanic force every bit well every bit the one-half time of force disuse from ninety% of maximum tetanic strength was slightly longer in RNR muscle than WT muscle (north = 22 in WT and n = 28 in RNR, unpaired T exam). C. Time course of equatorial intensity ratio during activation and relaxation. D. Half time of intensity ratio decay was longer in RNR muscle than in WT muscle (n = 9 in WT and n = 13 in RNR, unpaired T test).

Effigy 5.. Myosin head rearrangement during relaxation from WT and RNR muscle.

A. Normalized South_M3 recovery every bit function of time afterward the end of stimulation from WT (blackness) and RNR (blueish) muscle. B. Normalized S_M6 recovery as function of time later the end of stimulation from WT (black) and RNR (blue) muscle. C. Normalized MLL1 intensity equally part of time after the end of stimulation. D. The fourth dimension for recovery of MLL1 intensity after the end of stimulation for a tetanic contraction (n = 22 in WT and n = 28 in RNR, unpaired T test).

Figure vi.. Myosin head disposition in resting RNR and WT muscle

The resting equatorial intensity ratio (A) (northward = 29 in WT and n = 48 in RNR) and the average myosin caput radius (B) (n = 41 in WT and n = 48 in RNR) are larger in intact RNR mouse muscle than intact WT musculus indicating that the myosin heads are closer to actin in RNR muscle (unpaired T test). C. The resting lattice spacing are likewise larger in RNR mouse muscle than intact WT muscle (north = 72 in WT and n = 66 in RNR, unpaired T examination). The I₁₁/I₁₀ (D), S_M3 (E) and d_one,0 (F) from skinned WT muscle after treatment with relaxing solution containing v mM dATP instead of ATP (n = seven, paired T test).

Figure vii.. dATP induced concerted structural changes in the motor domain of myosin.

Snapshots taken from MD simulations of myosin II with ATP (grey ribbons, peak & heart) or dATP (blue ribbons, lesser) illustrate conformational changes that occurred to the motor domain as a event of dATP binding. dATP adopted multiple nonnative conformations, including an 'extended' conformation. Formation of the 'extended conformation' was associated with an opening of the nucleotide bounden pocket and an overall bending of the motor domain.

Effigy eight.. dATP induced concerted structural changes in the actin binding regions of myosin.

Snapshots taken from MD simulations of myosin II with ATP (grey ribbons, left) or dATP (blue ribbons, right) illustrate conformational changes that occurred to the actin biding regions as a effect of dATP binding. The dATP-induced opening of the nucleotide binding pocket and angle of the relay helix were accompanied past a reorganization of the actin binding residues and the formation of a unique conformation of the actin bounden pocket.

Figure 9.. dATP promoted exposure of positively charged residues in the actin binding surface of myosin.

These histograms report the probability densities of the solvent accessible expanse (SASA) of sets of actin binding residues in myosin. The left column reports the distributions for all side concatenation atoms, the right column reports the distribution for only the polar side chain atoms, the peak row reports the distributions for all actin bounden residues, and the lesser row reports the distributions for the positively charged actin binding residues. In all four conditions, dATP (bluish bars) shifted the SASA distributions to the correct of the ATP (black) distributions. This right shift corresponds to a consistent dATP-induced increase in the area of actin binding residues.

Effigy 10.. Loops two and 3 sampled extended conformations in the presence of dATP.

These histograms report the probability densities of the solvent accessible surface area (SASA) of all residues in loop ii (left) and loop 3 (right) in the ATP (black bars) and dATP (blueish bars) Doc simulations. Both loops sampled more extended conformations more ofttimes in the presence of dATP.

Effigy 11.. Contacts formed with the O-helix were nucleotide-dependent.

Contact maps highlight changes in side chain to side chain interactions observed in the X-ray construction (top), ATP MD simulations (heart), and dATP Md simulations (bottom). Each point in the graph corresponds to a side chain to side chain interaction between a residue in the O-helix (y-axis) and a residue in myosin (x-axis). The colour of the point corresponds to how oftentimes the contact was observed. Red arrows indicate regions of the structure where contacts shifted relative to the 10-ray construction

Effigy 12. Contacts formed with the relay helix were nucleotide-dependent.

Contact maps highlight changes in side concatenation to side chain interactions observed in the X-ray construction (top), ATP MD simulations (heart), and dATP Medico simulations (lesser). Each point in the graph corresponds to a side concatenation to side chain interaction between a residue in the relay helix (y-axis) and a residual in myosin (10-centrality). The color of the point corresponds to how ofttimes the contact was observed. Red arrows indicate regions of the structure where contacts shifted relative to the X-ray construction.

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