Contractile structure
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Contractile structure. Sarcomere structure Contractile filament composition Myosin (thick) Actin (thin) Anchors and attachments Z-disk Titin/nebulin. Basic Structure. M. Z. 1 um. Striated muscle. “Light” and “dark” bands under visible light Birefringence (uniaxial anisotropy).

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Contractile structure

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Contractile structure

Contractile structure

  • Sarcomere structure

  • Contractile filament composition

    • Myosin (thick)

    • Actin (thin)

  • Anchors and attachments

    • Z-disk

    • Titin/nebulin


Basic structure

Basic Structure

M

Z

1 um


Striated muscle

Striated muscle

  • “Light” and “dark” bands under visible light

  • Birefringence (uniaxial anisotropy)


Birefringence

Birefringence

  • Light intensity due to refraction

Focused over

Focused under

Light and dark

bands swap


Sources of striation

Sources of striation

  • Darker and lighter material bands

  • Wavy or crimped filaments

  • Bands of different refractive index

  • Disappears during contraction

  • Disappears by soaking in salt

Phase-contrast image of myofibril

Directly broken-up muscle

After extraction

(Hanson & Huxley, 1953)

5 um


Extractions

Extractions

  • High salt extracts of muscle coagulate over time

    • Myosin+actin+ATPActomyosin + ADP

    • Seems to be the active stuff of muscle

  • Flow-induced birefringence

  • Time-varying viscosity

  • Contraction results from

    • Polymerization (fibrin)

    • Rod shaped particles


X ray diffraction

X-ray Diffraction

  • 1-D diffraction

    • Constructive/destructive interference

    • ml = d sin q

  • Bragg diffraction

    • Reflection

    • nl = 2 d sin q

    • X-ray l~1-10Å

(Soren Pedersen)

d

q

d sin(q)


2 d diffraction

2-D diffraction

  • Composition of 1-D

    • Radial symmetry

    • Miller notation

Unit cell

Major diffraction along (100)

Major diffraction along (110)

NaCl crystal


Anisotropic crystals

Anisotropic crystals

  • 2-D image (still some symmetry)

  • Depends on illumination window

1,1

1,0

Myofilament image

Living

45 nm

3 axes of symmetry in point-illuminated image

H.E. Huxley’s image, near the axis of living muscle fiber shows strong reflection @ 45nm & weak reflection 22.5 nm. In rigor, these intensities reverse.

Rigor

22 nm

Huxley, 1953


Contractile structure

Relaxed muscle

Rigor muscle

“End-on” diffraction pattern

Intensity shifts toward thin filaments

Fourier transformed data

Huxley, 1953


2 d diffraction1

2-D diffraction

  • Point-source images are more complicated

    • 14.3 nm, 43.0 nm triple-symmetry

    • Mostly due to myosin

Magid & Reedy, 1980

Huxley 1953


High resolution tem

High resolution TEM

200 nm

Huxley, 1957


Transverse tem

Transverse TEM


Myosin molecule

Myosin molecule

  • Native hexamer

    • 2 heavy chains 180 kD

    • 4 light chains

  • Domains

    • Globular head

    • Helical tail

    • Tryptic fragments


S1 motor domain

S1 motor domain

  • ATPase

  • Actin binding

  • Sufficient for motility

Spudich lab movie


Contractile structure

Actin filament points into page

(Lower 50 kD)

Holmes et al 2003


Myosin filament

43 nm

14 nm

25 nm

Myosin filament

  • Triple helix

    • Diffraction symmetry

    • 14.5 nm repeat

  • Cryo-EM

Woodhead & al., 2005


Contractile structure

MHC1

MHC2

ELC1

ELC2

RLC1

RLC2


M line

M-line

  • Thick filaments anchored at M-line

    • Myomesin

    • Obscurin

Thick Filament

Myomesin

Titin


Actin

Actin

  • Disk shaped

  • Adenine nucleotide binding

    • ATPase activity

    • Nucleotide exchange

      • Promoted by Profilin

      • Inhibited by Cofilin

  • Filament formation

    • Barbed/Pointed end

    • Myosin S-1 “decoration”

    • ADP maturation


Actin filament polymerization

Actin filament polymerization

  • Asymmetric exchange of monomers

Myosin fragment


S1 decoration

Molecular

Model

S1 decoration

Actin filament

S-1 Fragments


Actin filament regulation

Actin filament regulation

  • Troponin/tropomyosin

  • Nebulin

  • CapZ (barbed)

Weak myosin binding

Strong myosin binding

Actin

Tropomyosin


Extra contractile support

Extra-contractile support

  • Extract contractile proteins

  • Intermediate filament ring around Z-disk

  • External scaffold (desmin)

  • Z-disk ghost

Wang & Ramirez-Mitchell, 1983


Z disk

Z-disk

  • Thin filament anchor

Structural models

Transverse TEM

Long TEM

Rowe, 1971

Luther, 2009


Z disk1

Z-disk

  • a-actinin

  • Titin

  • F-actin


Titin

Titin

  • Molecular ruler

    • 3-4 MD

    • 30,000 AA

  • Modular spring


Titin1

Titin

  • Modular spring

    • Fn repeats

    • Ig repeats

  • Kinase

Hoshijima 2006

Labeit & al 2003


Summary

Summary

  • Sarcomere

    • Z-I-A-I-Z

    • Interdigitating arrays of thick & thin filaments

  • Myosin motors

  • Actin rails

  • Z-disk anchors

  • Titin skeleton


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