Urry D.W. (Ed.) What Sustains Life? : Consilient Mechanisms for Protein-Based Machines and Materials

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8.5 The Myosin II Motor of Muscle Contraction, a Representative ATPase

435

N-terminal

domain

Y-axis

' • X-axis

Z-axis is out

the

plane

terminal

domain

Essential

light chain

FIGURE 8.49. Stereo view (cross-eye) in space-filling

representation of the scallop muscle cross-bridge

(SI) with the essential light chain in ribbon repre-

sentation, without the regulatory light chain shown

and with neutral residues light gray, aromatic

residues black, other hydrophobic residues gray, and

charged residues white. The coordinate system places

the foot of the lever arm pointing in the direction of

the myosin filament from which it arises. This pro-

jection of the XY plane directly observes the near

90° kink as the a-heUcal lever arm connects the

myosin filament to the globular portion of the cross-

bridge. (A) Near-rigor state. (Prepared using the

crystallographic results of Himmel et al.^ as obtained

from the Protein Data Bank, Structure File 1KK7.)

(B) ATP analogue state (ADP-BeFx). (Prepared

using the crystallographic results of Himmel et al.^ as

obtained from the Protein Data Bank, Structure File

1KK8.)

436

8. Consilient Mechanisms for Protein-based Machines of Biology

Z-axis is out

the

plane

X-axis

AN-terminal

vJ domain

Essentia]

light chai

FIGURE 8.50. Same projection as in Figure 8.49

except that the cross-eye stereo view is of a ribbon

representation of the scallop muscle cross-bridge

(SI) with the essential Ught chain included but no

regulatory Ught chain and with neutral residues Ught

gray, aromatic residues black, other hydrophobic

residues gray, and charged residues

white.

The coor-

dinate system is chosen with the foot of the lever arm

pointing in the direction of the myosin filament from

which it arises. This projection at the XY plane

X-axis

directly shows, as unchanged between the two states,

the 90° kink as the a-heUcal lever arm connects the

myosin filament to the globular portion of the cross-

bridge. See text for discussion. (A) Near-rigor state.

(Prepared using the crystallographic results of

Himmel et al.^ as obtained from the Protein Data

Bank, Structure File 1KK7.) (B) ATP state with the

ATP analogue (ADP-BeFx). (Prepared using the

crystallographic results of Himmel et al.^ as obtained

from the Protein Data Bank, Structure File 1KK8.)

8.5 The Myosin II Motor of Muscle Contraction, a Representative ATPase

437

8.5.4.3.3

View of the Cross-bridge in the XZ

Plane Demonstrates Major Change in

Relationship Between the Head of the Lever

Arm and the Amino-terminal Domain

In the space-filling representation in Figure

8.51,

the perspective is essentially looking up

the lever arm from the foot to its head. In

this view of the projection in the XZ plane,

the major discernible structural rearrangement

involves the amino-terminal domain. The same

perspective is given in backbone representation

in Figure 8.52. To get the perspectives the same

for both states, the Z-axis is sighted directly

N-terminal

domain

X-axis

Y-axis'-'

into the plane

N-terminal

domain

FIGURE

8.51.

Stereo view in space-filling representa-

tion of scallop cross-bridge (SI) without light chains

but with neutral residues in light gray, aromatics in

black, other hydrophobics in gray, and charged

residues in white. This perspective of the XZ plane,

looking in the negative Z-direction, provides the best

view of the movement of the amino-terminal domain

that is positioned as a flap over the head of the lever

arm in A and moves to be a free standing pedicle in

The best view of this movement is shown in the

same perspective but in backbone representation as

in Figure 8.52. (A) Near-rigor state. (Prepared using

the crystallographic results of Himmel et al.^ as

obtained from the Protein Data Bank, Structure File

1KK7.) (B) ATP analogue state (ADP-BeFx). (Pre-

pared using the crystallographic results of Himmel et

al.^

as obtained from the Protein Data Bank, Struc-

ture File 1KK8.)

438

8. Consilient Mechanisms for Protein-based Machines of Biology

N-terminal domain

folded over the head

the

lever arm

—> X-axis

Y-axis ^ into the plane

FIGURE 8.52. The same perspective for the stereo

view (cross-eye) of scallop cross-bridge (SI) as

shown in Figure 8.51, except that it is in backbone

representation and thereby allows better delineation

of domain movements. In A the amino-terminal

domain is shown as a flap over the head of the lever

arm in the near-rigor state, whereas it separates as a

free-standing pedicle in

the ATP

state.

The

amino-

terminal domain movement also becomes apparent

by the changing G53 location at its leading edge.

N-terminal domain

pedicle separates

from upper lever arm

on phosphorylation.

X-axis

Another interesting feature in this backbone repre-

sentation of the XZ plane perspective is the chain

segment that appears stretched in A but relaxed in

(A) Near-rigor state. (Prepared using the crystal-

lographic results of Himmel et al.^ as obtained from

the Protein Data Bank, Structure File 1KK7.) (B)

ATP analogue state (ADP-BeFx). (Prepared using

the crystallographic results of Himmel et al.^ as

obtained from the Protein Data Bank, Structure File

1KK8.)

along the a-helix of the low^er part of the lever

arm from residue 825 to residue 800. From this

viewpoint the rotation of the 50kDa upper

domain is apparent from comparison of the

reorientation of the major a-helix of the upper

domain. This clockwise rotation of the 50kDa

upper domain, as seen from above, on going

from the ATP state to the near-rigor state,

appears to result from the amino-terminal

domain movement. In what follows in Figures

8.5 The Myosin II Motor of Muscle Contraction, a Representative ATPase

439

8.53 and 8.54, the hydrophobic consiUent mech-

anism provides an explanation of the physical

basis for this movement.

Another feature of this perspective involves

the apparent stretching of a chain segment, as

labeled in Figure 8.52. The elastic force result-

ing from such an extension derives from the

elastic consihent mechanism. As will be argued

below, hydrophobic association gives rise to

the development of a functional elastic force.

Such a perspective may become a classic

demonstration in protein-based machines, as it

has been seen in Complex III of the electron

transport chain and in the repulsive aspect of

the comprehensive hydrophobic effect causing

an elastic deformation during functioning of

ATP synthase/Fi-ATPase.

8.5.4.4 Hydrophobic

Association/Dissociation of the Head of

the Lever Arm and the Amino-terminal

Domain

Figure 8.53 shows stereo views of the scallop

muscle cross-bridge, absent the Ught

chains,

that

were oriented to better characterize the nature

of the association (in the near-rigor state) and

the dissociation (in the ATP bound state)

involving the head of the lever arm and the

amino-terminal domain. The stereo views help

to show that the association in Figure 8.53A,C

is hydrophobic, whereas the proximal surfaces

of the head of the lever arm and amino-

terminal domain in Figure 8.53B,D are domi-

nated by charged residues.

Enlarged views of the relationship between

the amino-terminal domain and the head of the

lever arm, shown in stereo in Figure 8.53, are

given in Figure 8.54 for the purpose of better

seeing the interacting residues in the associated

near-rigor state. On the left side of the figure

the association is hydrophobic, with, as seen in

the stereo view of Figures 8.53A, the lower

charged part between the domains having

twisted sUghtly out of

co-planarity.

On the other

hand, the ATP analogue state is dissociated

with a preponderance of charged (white)

residues separating the two surfaces.

The explanation for the structural transition

between the near-rigor and ATP bound states

is given in section

8.5.4.7

(with the assistance

of Figures 8.58 and 8.59). First, however, two

issues require discussion before this pair of

states can be considered models for producing

the motion of muscle contraction. One issue

involves a closer examination of the stretched/

relaxed chain segment of Figure 8.52 in order

to see if this chain segment can provide an

example of elastic force development that

results from hydrophobic association demon-

strated in Figures 8.53 and 8.54. The pair of

structures to be of greater significance should

provide an understanding of force develop-

ment, as required to explain isometric con-

traction and to understand energy-efficient

production of motion. The second issue is how

effectively the two structures demonstrate the

displacement required for the sliding filament

mechanism of muscle contraction.

8.5.4.5 Evidence for Elastic Force

Development Resulting from

Hydrophobic Association

8.5.4.5.1

Stretching the Relay Loop on

Hydrophobic Association of the Head of the

Lever Arm and the Amino-terminal Domain

The suggestion of a stretched chain segment is

shown in Figure

8.52.

An optimized perspective

for considering this potential source of elastic

force development is given in Figure 8.55. The

estimate of length change uses the change of

ratios of the relay loop segment I505-D511 to

the relay helix segment Y500-F472 that accom-

panies the change of states at the nucleotide

binding

site.

There appears to be an extension of

as much as

40%

accompanying the hydrophobic

association resulting from loss of phosphate (or

equivalent) from the nucleotide binding site.

8.5.4.5.2

Presence of a Number of Flexible

Loops for Potential Storage of Elastic Force

By the elastic consihent mechanism the

extension of single flexible loops causes an

increase in the elastic force. It appears that one

such loop has been identified in Figure 8.55.

Rayment et al.^^"^ note a number of flexible

loops in the myosin cross-bridge. Each of

these becomes a candidate for elastic force

440

8. Consilient Mechanisms for Protein-based Machines of Biology

1 -..-i

^B^-i-

«.'C«^

^TP site^

Pin clef^jJFte

riln^^^HII^^B-4k'

«>.

Rli^^^BHki'"^

KrrZpk

i^T^^^jh

FIGURE 8.53. Cross-eye stereo views in space-filling

representation of scallop muscle cross-bridge (SI),

without essential and regulatory fight chains but with

cross-bridge oriented to better show the changing

relationship between the lever arm and the amino-

terminal domain.

facilitate observation of hydro-

phobic domains and their association, neutral

residues are fight gray, aromatic residues are black,

other hydrophobic residues are gray, and charged

residues are white. (A,C) Near-rigor state showing

hydrophobic association between the amino-

terminal domain and the upper part of the lever arm

with the view taken approximately along the axis of

the myosin filament. A shows the side of the

nucleotide binding site referred to as the front side,

whereas C shows the opposite (back) side. (Prepared

using the crystallographic results of Himmel et al.^ as

obtained from the Protein Data Bank, Structure Ffie

1KK7.) (B,D) ATP analogue state (ADP-BeFx)

showing hydrophobic dissociation of the amino-

terminal domain and the upper part of lever arm. B

shows the side of the nucleotide binding site referred

to as the front side, whereas D shows the opposite

(back)

side.

View now more nearly perpendicular to

the myosin filament, that is, the lever arm rotates

with an untwisting on hydrophobic dissociation.

(Prepared using the crystallographic results of

Himmel et al.^ as obtained from the Protein Data

Bank, Structure File 1KK8.)

8.5 The Myosin II Motor of Muscle Contraction, a Representative ATPase

441

FIGURE 8.54. Scallop muscle cross-bridge (SI):

Enlarged non-stereo views and orientations of those

in Figure 8.53 to characterize the nature of the

changing relationship between the amino-terminal

domain and the head of the lever arm with charged

residues in white, aromatic residues in black, other

hydrophobic residues in gray, and neutral residues in

light gray. (A,C) Near-rigor state with nucleotide

binding site (front side) at the upper left and the

opposite (back side) at the lower left. The shading of

the associating residues demonstrates hydrophobic

^1^

ll»^

t&A.^

L'-i»;

association. (Prepared using the crystallographic

results of Himmel et al.^ as obtained from the

Protein Data Bank, Structure File 1KK7.) (B,D)

The ATP analogue (ADP-BeFx) bound state at the

upper right (front side) and lower right (back side)

showing hydrophobic dissociation due to the pre-

ponderance of charged residues separated by a wide

aqueous corridor. (Prepared using the crystallo-

graphic results of Himmel et al.^ as obtained from

the Protein Data Bank, Structure File 1KK8.)

442

8. Consilient Mechanisms for Protein-based Machines of Biology

rF472

Stretched

chain

•Y500

.F472

¥500

FIGURE 8.55. Expanded stereo view (cross-eye) of

scallop muscle cross-bridge (SI) in ribbon represen-

tation to estimate length change of a portion of the

relay

loop.

The

ratio of the length of

I505-D511

loop

segment to that of the Y500-F474 segment of the

relay

heUx,

when comparing the two states, indicates

as much as a 40% extension on hydrophobic associ-

ation attending loss of phosphates. (A) Near-rigor

state without nucleotide in the binding site. (Pre-

pared using the crystallographic results of Himmel

et al.^ as obtained from the Protein Data Bank,

Structure File 1KK7.) (B) ATP state using the ATP

analogue ADP-BeFx. (Prepared using the crystallo-

graphic results of Himmel et al.^ as obtained from

the Protein Data Bank, Structure File 1KK8.)

development on hydrophobic association

attending dephosphorylation, as most readily

evaluated during isometric contraction, that is,

force development at fixed length. The increase

in elastic force on extending a single chain, as

determined from single-chain force-extension

studies on elastic-contractile model protein,

suggest that several chains would be stretched

during the hydrophobic association of muscle

contraction to achieve adequate force levels.

8.5 The Myosin II Motor of Muscle Contraction, a Representative ATPase 443

8.5.4.6

Use ofActin Binding Site on the

Myosin Cross-bridge as a Reference

to Evaluate Changes in Lever

Arm Orientation

To consider a pair of states of the myosin cross-

bridge (such as the near-rigor and ADP-BeFx

states being analyzed above) as relevant to the

mechanism of muscle contraction, there should

be evidence of lever arm displacement. No sig-

nificant displacement was in evidence in the

perspectives utilized in Figures 8.47 through

8.53.

The perspectives in Figures 8.56 and 8.57

use the actin binding site on the myosin cross-

bridge as a reference for evaluating changes in

orientation of the lever arm that w^ould result

from dephosphorylation.

The motion of the lever arm appears best

described as a tucking under the amino-termi-

nal domain due to hydrophobic association. The

N-terminal domain

with lever arm tucked

under it by means of

drophobic association

'^X-axis

^ movement

along actin

filament

N-terminal domain

separated from head

of the lever arm

FIGURE 8.56. Stereo view in space-filling representa-

tion of a scallop cross-bridge (SI) view of the actin

binding site on myosin cross-bridge, with continuity

of the cleft from the ADP-BeFx binding site to the

actin binding site that dissociates on ATP binding (A)

Near-rigor state without nucleotide in the binding

site.

(Prepared using the crystallographic results of

Himmel et al.^ as obtained from the Protein Data

Bank, Structure File 1KK7.) (B) ATP state using the

ATP analogue ADP-BeFx- (Prepared using the crys-

tallographic results of Himmel et

al.^

obtained from

the Protein Data Bank, Structure File 1KK8.)

444

8. Consilient Mechanisms for Protein-based Machines of Biology

FIGURE 8.57. Stereo view of scallop muscle cross-

bridge (SI) in backbone representation from the

perspective of the actin binding site on the myosin

cross-bridge. (A) Near-rigor state without nucleotide

in the binding site. (Prepared using the crystallo-

graphic results of Himmel et al.^ as obtained from

the Protein Data Bank, Structure File 1KK7.) (B)

ATP state using the ATP analogue ADP-BeFx. (Pre-

pared using the crystallographic results of Himmel et

N-terminal domain

with lever arm tucked

under it by means of

hydropliobic association

'^X-axis

^ movement

along actin

filament

ATP-analogue

Y-Pj

location

N-terminal domain

separated from head

the

lever arm

al.^

as obtained from the Protein Data Bank, Struc-

ture File 1KK8.) On going to the near-rigor state, the

amino-terminal domain moves sUghtly forward as

the lever arm rotates about 90° and tucks its head

under the amino-terminal domain by means of a

hydrophobic association to provide for the major

motion of the powerstroke. On ATP binding this

hydrophobic association is lost due to apolar-polar

repulsion from phosphate in the active site.

hydrophobic association would be most effec-

tively disrupted by the ADP-Pi occupancy state

of the ATP binding site, and the hydrophobic

association would most effectively reform on

dephosphorylation. Importantly, the displace-

ment of the lever arm is as required for move-

ment along the actin filament. Also, the tucking

of the head of the lever arm accompanied by a

twist would stretch the relay loop of Figure 8.55

in the associated development of an elastic

force. These are as required for achieving the

motion of muscle contraction. What follows

immediately below is a clarification of the force

that would bring about the observed changes.