Myocardial proteins/contraction cycle Flashcards
Ultrastructure of cardiac muscle
- Myo¢ contain numerous myofibrils
- Myofibrils = longitudinally repeating sarcomeres
Define sarcomere
from Z line to Z line, 1.6-2.2um
o Thin filament: Z line to center
Form I band
o Thick filament: center (M line) => Z line
Form H band
o Overlap of filaments = darker zone = A band
Structure of thick filaments
MYOSIN
* Tail: confer stability
o Heavy chains form the A helical tail
o Light chains: link tail => heads
4 lights chains/ bilobed head
2 types: MLC-1 and MLC-2
* 2 heads: 2 distinct sites to interact w actin and hydrolyze ATP
o S1 subfragment: head
ATP binding => conformational change
o S2 subfragment: neck connecting the head to filament
- Myosin molecules form myosin filaments: 300-400 myosin heads
Myosine isoenzymes
2 isoforms of heavy chains = Aand B
o V1: 2 A chains = high ATPase activity => fast muscle contraction
o V2: AB heterodimer
o V3: 2 B chains = low ATPase activity => slow muscle contraction
Primarily in normal adult myocardium
Regulation of myosin ATP ase
Short term: incr [Ca2+] => incr myosin ATPase activity => V1
Long term: gene changes with CHf, hypertrophy -> V3
Role of myosin binding protein C
o Prevent interaction of actin and myosin decr [Ca2+]
o Stabilize S2 subfragment
Thin filaments structure
- 2 strains of fibrous actin => double A helix filament
o G actin monomere (multiple globular units) that assemble into filament (F-actin) at physio [ATP/Mg]
o Interact w globular head of myosin => form cross bridges
What are the regulatory proteins around contractile proteins
Ca2+ dependent regulation
o Troponin: globular prot
TnC: binds Ca2+ => conformational changes in tropomyosin
TnI: inhibit Mg stimulated actomyosin ATPase
* incr[Ca2+] => firmly bind to TnC (and not to actin) to allow contraction
TnT: attach troponin to tropomyosin
* incr Ca2+ => activate actomyosin ATPase
o Tropomyosin: double strand A helical chain
Lie in 2 major groove along actin filament in diastole
Titin role
- Link the myosin molecule to the Z line and stabilize filaments
- Provides elasticity: molecular spring modulating systole and diastole
o Expand and contract as sarcomere stretch
o incr systolic contraction in response to stretch and deformation - Role in transducing stretch signal into growth
o Interact w 2 prot of Z disc
Steps of excitation-contraction coupling
Resting state: tropomyosin block myosin binding sites on actin
a) Ca2+ bind to TnC => displace TnI => conformational change of tropomyosin
b) Exposition of actin sites => myosin heads binding sites
a. decr [ATP] => high affinity (severe hypoxia, ischemia, death)
b. Need ATP + Ca2+
c) ATP cleaved by myosin ATPase => ADP + Pi => bind myosin + actin => cross bridge
a. Formation of strong binding state
b. E derived from rx used to generate movement (ratchet like)
c. Rigor state
d) Release ADP + Pi => bind new ATP => myosin conformation change => dissociate actin/myosin
a. Conformation change: strong binding state to weak binding state until cycle starts again
Sarcomere shortening depend on
Cycle repeated to have significant sarcomere shortening
* If incr [Ca2+] => tropomyosin block active sites of actin
* Contraction force and rate of shortening => related to # cross bridges + [Ca2+]
* Cycling rate is dependent on the rate of attachment (F) and detachment (G) of the cross bridges
Control of contractile cycle by Ca2+
incr force development
* incr activation of cross bridges by Ca2+
o Binding Ca2+ to TnC
Molecular signal in actin filament => incr rate of cross bridges
TnC bind TnI => relieve inhibition
incr [Ca2+] => incr Ca2+ binding to TnC =>incr # cross bridges
o B adrenergic stimulation: cAMP => PKA
incr lusitropy (see below): phospholamban Pi + decr [Ca2+) will incr TnC dissociation
* Pi or dPi of contractile proteins (see below)
* Starling mechanism
Intracell Ca2+ normal
- Intra¢ [Ca2+] > extra¢ [Ca2+] => large chemical gradient for Ca2+ entry
o Ca2+ transient: cyclical variation in cytosolic [Ca2+]
incr by B adrenergic stimulation
Sarcolemmal channels important for excitation contraction coupling
L type channels Ca2+ channels
Na+/Ca2+ exchanger
Na+/K+ ATPase
Ca2+ pump
Structure of L type Ca2+ channels
- Structure: 5 subunits
o A1 => active unit, pore to enter Ca2+
o 4 repeated units containing 5 hydrophobic helices + 1 charged hydrophilic helix
Role of L type ca2+ channel
- Will sense change in voltage during depolarization in phase 2 of AP
- Opening => small amount of Ca2+ enters => stimulate release of larger amounts
Structure of SR and important channels
- Membrane limited structure intracell surrounding myofibrils
SERCA Ca2+ ATP ase pump
Ca2+ release channels RYR
Functions of SR
o Actively pump Ca2+ in its lumen during diastole
o Store Ca2+ in diastole
o Release Ca2+ in systole
incr with incr [Ca2+] triggering or incr amount of Ca2+ in SR
* incr # of channels or incr amount of Ca2+ released
Controlled by local [Ca2+]
Role of SERCA channels
o SERCA-2a is the isoform present in cardiac myo¢
Bind Ca2+ and ATP
o Activated by incr intra[Ca2+] => actively pump Ca2+ into SR lumen in diastole
Remove 75% of Ca2+
1ATP => moves 2 Ca2+ into SR
Regulation of SERCA channels
o Inhibited by phospholamban (monomer) Pi relieve inhibition
B adrenergic stimulation: cAMP => PKA
incr [Ca2+] promote Pi at 2nd site => incr rate of relaxation and Ca2+ stores
Structure Ca2+ release channels Ryanodine R
4 ryanodine R protein to form 1 RyR
Control of RyR
B adrenergic stimulation => PKA Pi
* inrc rate of incr/decr of intra¢ [Ca2+]
* Pi RyR => incr rate of Ca2+ release
* Pi phospholamban => incr SR Ca2+
A adrenergic stimulation => IP3
* incr Ca2+ release via IP3 R on RyR
Caffeine: incr probability of opening of RyR => can empty SR
o Closure: 3 possibilities
Continuing incr in local [Ca2+] at supraoptimal levels
Local decr or SR Ca2+
Stochastic attrition: spontaneous decay of active clusters due to random channel closure => all L type + release channels close at same time => decr [Ca2+]
o Ca2+ sparks: small amount of Ca2+ locally/spontaneously released w/o L type opening
Not enough to induce depol
Role of RyR
Stimulated by Ca2+ entering ¢ by L type channels
Conduit for rapid Ca2+ release from SR in systole
Ca2+ regulatory proteins/factors
- Calmodulin (CaM): regulatory protein
o Activated w Ca2+ binding - Calcineurin: CaM activated growth factor => sustained incr [Ca2+] => promote hypertrophy
- Calsequestrin: prot binding Ca2+ w/i SR (strong – charge)
o Allow rapid Ca2+ mvt
o Calreticulin is another storage protein similar in structure/fct
o Low affinity, high capacity - Phospholamban: on SR membrane => inhibit Ca2+ pump in dePi state
Structure of transverse tubular system (T system)
- Continuous w ¢ membrane, in close association w SR
- T tubule + junctional SR (terminal section) => form triads
o Foot proteins = 7-29 Ca2+ release channels close to 1 Ca2+ L type channels
Arrival of Ca2+ ions from L type channels => conformational change in foot => open release channels
Na+/Ca2+ exchanger role
3Na+ in for 1Ca2+ out
o Function: efflux of Ca2+ in diastole (25% of Ca2+ released by SR)
Activated by ↑ Ca2+ and ATP
Inactivated by Na+
o Can move Ca2+ both direction depending on [Na+]
o Transient Ca2+ influx can occur at depol state after Na+ channel opening => Na+ accumulate close to channels => transient activation of exchanger in reverse direction => incr Ca2+ => incr contraction
Na+/K+ ATPase role
maintain low [Na+] to allow Ca2+ efflux
o Activated by incr [Ca2+], ATP
o Inactivated by incr [Na+]
Ca2+ pump role
Ca2+ efflux w ATP
o Respond to incr [Ca2+] in diastole to maintain Ca2+ homeostasis
What determines contraction force
of active cross bridges => determine contraction force
* Normal heart: Ca2+ released for 10-25% myofilament activation
Cellular regulation of myocardial force generation
a) Amount of Ca2+ entering through L type channels
b) Amount of Ca2+ stored in SR
c) Myofilaments proteins => phosphorylation
d) Starling’s law
e) ATP availability: deficiency impairs performance
How amount of Ca2+ going into L type channels influence force of contraction
- incr activity with:
o B adrenergic stimulation: cAMP prot kinase phosphorylation
incr release by RyR
o Calmomodulin mediated prot kinase => activate cGMP
Respond to transient incr in [Ca2+] from hormones, NT, depol - incr activity with acidosis => slow channel
how amount of Ca2+ stored into SR influence force of contraction
- Autoregulation:
o incr SR [Ca2+] => incr RyR release => decr Ca2+ entry and incr efflux
o decr SR [Ca2+] => incr rapid refilling => SERCA activation - B adrenergic stimulation => G prot coupled => Gi or Gs
o Gs: activate AC => cAMP stimulat cAMP dependent prot kin => phospholamban phosphorylation => incr Ca2+ pump activity
How myofilament protein phosphorylation influence force of contraction
- TnI Pi: by cAMP dependent prot kin
o decr affinity of TnC for Ca2+ => incr relaxation by incr rate of Ca2+ dissociation - Myosin binding prot C Pi
- Myosin light chains Pi: calmomodulin kin, myosin light chain kin, PCK
o incr responsiveness to Ca2+
o incr force generation at low Ca2+ - TnT isoforms will change
o Myofibrillar ATPase activity
o Sensitivity of myofilaments to Ca2+
o Sarcomere length dependance of myofilament sensitivity to Ca2+
How starling laws influence force of contraction
incr contraction force with incr sarcomere length
* incr SV w incr end diastolic volume/pressure
* incr affinity of TnC for Ca2+ as sarcomere length incr => incr cross bridge #R
Role of mitochondria in intracell ca2+ control
*Mitochondria play no role in beat to beat control of intra Ca2+
* ↑ [Ca2+] w/ increase in intracell cytosolic Ca2+ → may stimulate citric acid and produce more ATP
* Buffer by storing excess Ca2+
Effect of CHF on pumps
- decr activity of SERCA + incr activity of Na+/Ca2+ exchanger => decr Ca2+ SR stores => decr contractility
- HyperPi of RyR => incr Ca2+ leaks
- decr # or mRNAs for RyR, SERCA and phospholamban
Effect of arrhythmias on intra cell Ca2+
Ca2+ overload
* 2nd to myocardial ischemia, reperfusion, incr catecholamine stimulation
* incr O2 demand by mitochondria
o Attempt to buffer incr intra¢ [Ca2+] => mitochondrial Ca2+ overload => ATP waste => incr O2 needs
* Stimulation of phospholipase enzymes => incr membrane breakdown
* Can cause excess sustained contraction => contracture
* Excess Ca2+ cycling in/out of SR => substrate for arrhythmias
