Scenario 5

1
Scenario 5
A-bands stay at centre of sarcomeres
2
Scenario 5
Hierarchical organization of striated
muscle Muscle is a tissue, i.e. population of
cells The cells are known as muscle fibres.
(Aquire highly elongated shape by end-to-end
fusion of ordinary-shaped cells, myoblasts) The
cells contain myofibrils, which are
organelles, (just as are mitochondria, except no
membranes) Sarcomeres are repeated structural
unit of myofibrils and contain several types of
filaments
3
Scenario 5
Thin filaments are microfilaments, assembled
during development from G-actin as in Scenario 4,
but no longer dynamic, and of accurately fixed
length
4
Scenario 5
Thin filaments are microfilaments, assembled
during development from G-actin, as in Scenario
4, but no longer dynamic, and of accurately fixed
length Also contain tropomyosin and troponin,
which together confer calcium-dependence on
contraction Troponin C is muscle isoform of
calmodulin
5
Scenario 5
Thick filaments are bipolar filaments assembled
during development by a precise aggregation of
tails of the motor protein myosin II Two-stages
of interaction 1) Two myosin heavy chains (MHC)
associate by forming coiled - coil of their
a-helices (heptad repeat). This is one myosin
molecule.
6
Scenario 5
Thick filaments are bipolar filaments assembled
during development by a precise aggregation of
tails of the motor protein myosin II. Two-stages
of interaction 1) Two myosin heavy chains (MHC)
associate by forming coiled - coil of their
a-helices (sequence with heptad repeat). This is
one myosin molecule. 2) Molecules associate,
largely by electrostatic interaction (opposite
charges attract).
7
Scenario 5
8
Scenario 5
Thick filaments continued Two-stages of
interaction 1) Two myosin heavy chains (MHC)
associate by forming a coiled - coil of their
a-helices (heptad repeat). This is one myosin
molecule 2) Molecules associate, largely by
electrostatic interaction (opposite charges
attract). (Myosin molecules are soluble in high
salt, 0.6M KCl, and on dilution to 0.1M,
filaments self- assemble)
9
Scenario 5
Myosin superfamily As kinesin, the myosin motor
domain, recognizable by its amino acid sequence,
occurs in a large number of different molecules,
with varied functions. At least 17
families. Most are plus end- (barbed end)
directed motors. Many do not form filaments, and
are involved in, e.g. vesicle transport on actin.
Cf kinesin on microtubules.
10
Scenario 5
Myosin about mechanism Historically, studied as
muscle actomyosin (gel contracts on adding ATP-Mg
2 Ca 2)
11
Scenario 5
Myosin about mechanism Historically, studied as
muscle actomyosin (gel contracts on adding ATP-Mg
2 Ca 2) Motor assay reveals activity in S1
motor domain of myosin from muscle, and activity
of unconventional myosins. (S1 subfragment
1, cleaved from native molecule by proteolysis).
12
Scenario 5
Myosin about mechanism Conformation change on
binding or release of ligand is a general
property of proteins.
13
Scenario 5
Myosin about mechanism Conformation change on
binding or release of ligand is a general
property of proteins. Compare induced fit
model of enzyme-substrate binding
14
Scenario 5
Myosin about mechanism Conformation change on
binding or release of ligand is a general
property of proteins. Myosin is believed to use
a lever arm to magnify such a change. There
has to be some sort of linkage between ATP
hydrolysis, and binding of motor to actin. Rigor
mortis When ATP in cells has run down, muscles
stiffen because motor domains bind strongly to
actin. (cf organelle binding via kinesin in
AMPPNP)
15
Scenario 5
16
Scenario 5

• Myosin about mechanism
• Relation between enzymic and mechanical events
• presence of ATP, but without Ca 2 activation
  of thin filament, muscle is relaxed. Minimal
  interaction with actin.
• Even in absence of actin, ATP is hydrolysed.
  But this is not a complete enzymic cycle, because
  products are not released.

17
Scenario 5

• Myosin about mechanism
• Relation between enzymic and mechanical events
• Release of the phosphate allows binding to
  actin (when thin filament is activated)
• Release of ADP causes the mechanically
  important conformation change power stroke
• Further cycling requires ATP, hence rigor state
  in its absence

18
Scenario 5
19
Scenario 5

• Muscle length/tension relationship
• Force which can be generated is proportional
  to number of myosin motor domains with access to
  actin
• In physiological range (A-B on diagram) is
  proportional to extent of overlap of thick and
  thin filaments
• This is potentially unstable, because if an
  A-band moves slightly off centre, it experiences
  greater force in the direction of the error.
• Therefore need a centring mechanism - elastic
  protein titin

20
Scenario 5

• Titin
• Longest single polypeptide known. gt 3
  Megadaltons 3 x 106
• Characteristic of sarcomeric muscles. i.e
  worm, fly, man, not yeast!
• One end links to Z-disc, the other to centre
  of A band (i.e. of myosin bipolar filaments)
• gt300 domains arranged in tandem along length.
  Domains are immunoglobulin-like (IgG) or FnIII

21
Scenario 5

• Titin
• Longest single polypeptide known. gt 3
  Megadaltons 3 x 106
• Characteristic of sarcomeric muscles. i.e
  worm, fly, man, not yeast!
• One end links to Z-disc, the other to centre
  of A band (i.e. of myosin bipolar filaments)
• gt300 domains arranged in tandem along length.
  Domains are immunoglobulin-like (IgG) or Fn III
  (Fibronectin Type 3).
• IgG domains of I band part denature reversibly
  under tension

22
Scenario 5
A-bands stay at centre of sarcomeres
23
Scenario 5