Chapter 7 Proteins Function
1,Hemoglobin is a multisubunit allosteric (变构 )
protein that carries O2 in erythrocyte.
1.1 Hemoglobin is a well-studied and well-understood
protein.
1.1.1 It was one of the first proteins to have its
molecular mass accurately determined.
1.1.2 The first protein to be characterized by
ultracentrifuge.
1.1.3 The first protein to be associated with a
specific physiological function.
1.1.4 The first protein with a single amino
acid substitution being related to a genetic
disease (the beginning of molecular medicine).
1.1.5 The first multisubunit protein with
its detailed atomic structure determined by X-
ray crystallography.
1.1.6 The best understood allosteric
protein.
1.2 Determination of the atomic structure of
hemoglobin A (from normal adult) is very revealing.
1.2.1 The protein molecule exists as a a2b2
tetramer,
1.2.2 Each subunit has a structure strikingly
and unexpectedly similar to each other and to that
of myoglobin,indicating quite different amino acid
sequences can specify very similar 3-D structures.
1.2.3 Extensive interactions exist between the
unlike subunits through noncovalent interactions.
1.2.4 Quaternary structure changes markedly
when O2 binds,Crystals of deoxyhemoglobin shatter
(break) when exposed to O2.
1.2.5 O2 binds to the sixth coordination
position of the ferrous iron (as in myoglobin).
1.3 Hemoglobin is a much more intricate and sentient
(sensitive) molecule than is myoglobin.
1.3.1 The oxygen-binding (dissociation) curve of
hemoglobin is sigmoidal and that of myoglobin is
hyperbolic.
1.3.2 Myoglobin has a higher affinity for O2,
evolved for O2 storage.
1.3.3 Hemoglobin releases O2 efficiently at low
oxygen level tissues (thus evolved for O2 delivery),while
myoglobin does not.
1.3.4 Oxygen binding of hemoglobin shows
positive cooperativity,The binding of O2 (the ligand) at
one heme facilitates the binding of O2 at the other hemes
on the same tetramer (vice versa,unloading of oxygen at
one heme facilitates the unloading of oxygen at the
others),(Negative cooperativity refers to a decrease of
activity.)
1.3.5 Increasing concentrations of H+ (with a
decrease of pH) or CO2 lower the O2 affinity of
hemoglobin (H+ and CO2 has no effect on O2 affinity of
myoglobin),This is called Bohr effect,which helps the
release of O2 in the capillaries of actively metabolizing
tissues,
1.3.6 One molecule of 2,3-diphosphoglycerate (BPG)
binds to the central cavity of one tetramer of hemoglobin,
which lowers its O2 affinity.
1.3.7 BPG plays critical roles in the physiological
adaptation to the lower pO2 available at high altitude.
1.3.8 Fetal hemoglobin (HbF) binds BPG less
strongly than does hemoglobin A (adult) and consequently
has a higher oxygen affinity,
Identity
between
Hba and
Hbb,< 50%
Number of
identity AAs
among Mb,
Hba and
Hbb,27
kPa,kilopascal,1 atm = 760 mm Hg = 14.7 psi = 101.3 kPa.
binding sites occupied [L]
? = ------------------------------- = -------------
total binding sites [L] + Kd
? is a measurable quantity in experiment.
[L] is the free ligand concentration.
Hill plot,The ordinate = n log[L] - log Kd
Cooperativity,nH = 1,no; nH > 1,positive;nH < 1,negative.
Bohr’s effect and its molecular mechanism
Release of O2 in
peripheral tissues
Binding of O2 in lungs with
release of H+ and CO2.
1.4 Cooperativity is a particular case of an
allosteric effect
1.4.1 Allosteric effect refers to the
phenomenon in which a molecule (allosteric
effector) bound to one site on a protein causes a
conformational change in the protein such that
the activity of another site on the protein is
altered (increased or decreased).
1.4.2 H+,CO2,and BPG all show an
allosteric effect for the O2 binding process of
hemoglobin.
1.5 Two models have been proposed to explain the
allosteric regulation phenomena
1.5.1 The sequential model (proposed by
Daniel Koshland,Jr.) hypothesizes that the binding
of one ligand to one subunit changes the
conformation of that particular subunit from the T
state (with a low activity) to the R state (with a high
acitvity),a transition that increases the activity of
the other subunits for the ligand,
1.5.2 The sequential model can be analogized
to the tearing process of postage stamps,
1.5.3 The concerted model (proposed by Monod,
Wyman,and Changeux) hypothesizes that symmetry
is conserved in allosteric transitions (all subunits are
in the same conformation) and the binding of each
ligand increases the probability that all subunits in
that molecule are converted to the R-state (with a high
activity),All-or-none model.
1.5.4 The interplay between these different
ligand-binding sites is mediated primarily by changes
in quaternary structure,The contact region between
two subunits can serve as a switch that transmits
conformational changes from one subunit to another.
T,tense state,circle,less active;
R,relaxed state,square,more active
Concerted,all-or-none Sequential
1.5.5 The functional characteristics of an
allosteric protein are regulated by specific
molecules in its environment,In other words,in
the evolutionary transition from myoglobin to
hemoglobin,a macromolecule capable of
perceiving information from its environment has
emerged.
1.6 Sickle-cell anemia was found to be caused by a single
amino acid change in the b chain of hemoglobin molecules.
1.6.1 The hemoglobin molecule from sickle-cell
anemia patients (HbS) was found to have a higher pI value
(having more net positive charges).
1.6.2 Peptide fingerprinting (protease digestion +
electrophoresis + chromatography) of HbS and HbA (wt)
revealed that all but one of the peptide spots matched.
1.6.3 Amino acid sequencing revealed that HbS
contains Val instead of Glu at position 6 of the b chain!
1.6.4 DNA sequencing of the genes further revealed
that the codon of b6 changed from GAA(Glu) to GTA(Val).
1.6.5 The oxygen binding affinity and allosteric
properties of hemoglobin are virtually unaffected by this
change (the b6 is located at the surface of the protein).
1.6.5 High concentration of deoxygenated HbS
forms fiber precipitates,which sickles the red blood
cells,because the fiber formation is a highly concerted
reaction.
1.6.6 Presence of Val6 on the b subunits generates
a hydrophobic patch on the surface which
complements with another hydrophobic patch formed
only in deoxygenated HbS,thus generating the fiber
aggregates (a polymer of HbS).
1.6.7 Sickle cell trait (heterozygote) confers a
small but highly significant degree of protection against
the most lethal form of malaria (probably by
accelerating the destruction of infected erythrocytes,in
Africa).
1.6.8 Fetal DNA can be analyzed for the presence
of the HbS gene (prenatal DNA diagnosis).
1.7 Thalassemias are genetic disorders
characterized by defective synthesis of one or
more hemoglobin chains.
This can be caused by a missing gene,
impaired RNA synthesis or processing,
generation of grossly abnormal proteins.
Mutation and molecular interaction’s changes
2,Immunoglobulin superfamily members,found on cell surfaces
or secreted,are widely used for specific molecular recognition.
2.1.1 Class I MHC proteins bind and display
peptides derived from proteolytic degradation and
turnover of proteins that occurs randomly within the
cell,while Class II MHC detect peptides derived not
from cellular proteins but from external proteins
ingested by the cells.
2.1.2 Class I MHC occurs on the surface of
virtually all vertebrate cells,while Class II MHC is
found on the surfaces of a few specialized cells that
take foreign antigens,including macrophages and B
lymphocytes.
2.1 Major histocompatibility complex (MHC) detects
foreign protein antigens.
2.1.3 Class I MHC has up to 6 variants,while
each human is capable of producing up to 12 variants
of Class II MHC.
2.1.4 The complexes of Class I MHC and its
binding peptide are the targets of the T-cell receptors
of Tc cells in the cellular immune system,while those
for Class II MHC are the binding targets of the T-cell
receptors of Th cells.
2.1.5 Both Tc and Th cells undergo stringent
selection during the maturation process.
2.1.6 T cells patrol the infections foreign antigens,
and have life time of only a few days,Only a minority
of the T cells will trigger full immune response.
2.2 An immunoglobulin G (IgG) molecule (of ~150
kD) was found to contain two light and two heavy
chains,connecting to each other through disulfide
bonds.
2.2.1 Papain digestion convert the IgG
molecule into two Fab (antigen binding) and one Fc
(crystallize) fragments.
2.2.2 Each Fab fragment (containing one
complete light chain and half of a single heavy
chain) binds one molecule of antigen in a similar
manner to the original immunoglobulin molecule.
2.2.3 Each IgG molecule binds to two
molecules of antigens (thus called bivalent).
2.3 Complete amino acid sequence analysis of purified
myeloma patient’s immunoglobulins revealed strikingly
that the L and H chains consist of variable and constant
regions.
2.3.1 Residues 1 to 108 in the L chains are
relatively variable,and 109 to 204 relatively constant.
2.3.2 Residues 1 to 108 in the H chains are
variable,and 109 to 446 relatively constant.
2.3.3 Three segments in the L chain and three in
the H chain display far more variability than do others,
which are thus named as hypervariable segments (also
called complementary-determining regions,or CDRs,
because they determine antibody specificity).
2.3.4 Amino acid sequence analysis revealed that
the variable region of the light chain (VL) is similar in
sequence to that of the heavy chain (VH).
2.3.5 The constant region of each heavy chain
can divided into three parts (CH1,CH2,CH3) of
similar sequences.
2.3.6 The amino acid sequence of the constant
region of each light chain (CL) is similar to that of the
three parts in the constant region of the heavy chains.
Rodney Porter and Gerry Edelman were
awarded the Nobel Prize in 1972 in Medicine or
Physiology for their structure-function studies on the
antibody molecules.
2.4 X-ray crystallography studies revealed,strikingly,
that the immunoglobulin molecules are made of 12
structurally similar domains.
2.4.1 Each domain has a recurring structural
motif,called the immunoglobulin fold consisting of two
broad sheets of antiparallel b-strands joined by a
disulfide bond.
2.4.2 The CDRs of both VL and VH are located
in loops at one end of the sandwich made of the two b-
sheets that come together to form one antigen binding
site.
2.4.3 The immunoglobulin core serves as a
framework that allows almost indefinite variation of
the CDR loops,corresponding to various antigen
specificity of various antibodies.
2.5 T cell receptors,Class I and Class II Major
histocompatibility complex (MHC) proteins,and
intracellular adhesion molecules (ICAMs) all contain
domains similar to the immunoglobulin domains,thus
belonging to the immunoglobulin superfamily.
2.5.1 The immunoglobulin domains (folds) are
widely observed in these protein molecules,revealed by
sequence homology (~20% identity) and similar foldings
in some are confirmed by X-ray structure determination.
2.5.2 All these proteins are involved in molecular
recognition (antigen recognition by antibodies,TCR,and
MHC proteins; cell-cell interactions by ICAMs).
2.5.3 Most of the immunoglobulin superfamily
members are found on cell surfaces (e.g.,IgM,TCR,
MHC) or secreted (IgG).
3,ATP-driven conformational cycles lead to muscle
contraction.
3.1 Striated muscles contain overlapping arrays of
thick and thin filaments.
3.1.1 The thick filaments are primarily made of
myosin protein (肌球蛋白 ).
3.1.2 The thin filaments are primarily made of
actin (肌动蛋白 ),tropomyosin (原肌球蛋白 ),and the
troponin (肌钙蛋白 ) complex.
3.1.3 Thick and thin filaments slide past each
other in muscle contraction (a model proposed based
on X-ray,light microscope,and EM studies,fig.)
3.2 The force of muscle contraction arises from
the interplay of myosin,actin,and ATP.
3.2.1 Myosin consists of two globular
heads joined to a long a-helical coiled coil tail.
3.2.2 The myosin molecule can be cleaved
by trypsin into two partially functional
fragments,with one being able to form
filaments,and the other being ATPase and able
to bind actin.
3.2.3 Myosin molecules spontaneously
assemble into filaments in solutions of
physiological ionic strength and pH.
3.2.4 Actin molecules exist in monomer form (G-
actin) at low ionic strength and polymerize into a
fibrous form (F-actin,very similar to the thin filaments)
at physiological (higher) ionic strength.
3.2.5 Threads of actomyosin (肌动球蛋白 )
complex are formed when mixing actin and myosin in
solution.
3.2.6 Addition of ATP dissociates actomyosin into
actin and myosin.
3.2.7 The actomyosin threads contract when
immersed in a solution containing ATP,K+,and Mg2+,
whereas threads formed from myosin alone do not.
3.2.8 Myosin,being an ATPase,can be
regarded as a mechanoenzyme catalyzing the
conversion of chemical bond energy into
mechanical energy.
3.2.9 The ATPase activity of myosin is
markedly enhanced when it binds to the
polymerized form of actin (F-actin).
3.2.10 The hydrolysis of ATP drives the
cyclic association and dissociation of actin and
myosin,
3.3 The power stroke in contraction is driven by
conformational changes in the myosin head.
3.3.1 In resting muscle,the myosin heads,with
bound ADP and Pi,are unable to interact with the actin
units in thin filaments because of steric interference by
tropomyosin,a regulatory protein.
3.3.2 When muscle is stimulated,tropomyosin
shifts position,and the myosin head (with bound ADP
and Pi) reaches out from the thick filament and interact
with the actin units on thin filaments.
3.3.3 The binding of myosin-ADP-Pi to actin leads
to the release of ADP and Pi,which induces a major
conformational change in the myosin head pulling the
actin filament forward for about 100 Angstroms.
3.3.4 Subsequently,ATP binds to the
myosin head (myosin prefers binding to ATP than
to actin) and thus detaches it from actin.
3.3.5 Finally,the bound ATP is hydrolyzed
by the free myosin head,resetting it for the next
interaction with the thin filament.
3.3.6 The essence of the process is a cyclic
change both in the conformation of the myosin
head and its affinity for actin.
3.3.7 The control of protein-protein
interactions by bound nucleotides (ATP,GTP,etc)
is a recurring theme in biochemistry.
3.4 Troponin and tropomyosin mediate the
regulation of muscle contraction by Ca2+.
3.4.1 Ca2+ is released from sarcoplasmic
reticulum in muscle cells at the stimulation of a
nerve impulse.
3.4.2 Ca2+ controls muscle contraction by
an allosteric mechanism (through
conformational changes) in which the flow of
information is in the following order,Ca2+,the
troponin complex,tropomyosin,actin,myosin,
3.5 Other accessory proteins maintain the
architecture on the myofibril and provide it with
elasticity.
3.5.1 Springlike titin (肌联蛋白 ) molecules
(2.7 MD,or more than 25,000 amino acid
residues),the largest protein so far found in
nature,extend from the thick filaments to the Z
disc.
3.5.2 Nebulin,another large protein
molecule,is closely associated with the actin thin
filaments,and consists of almost entirely of a
repeating 35-amino-acid acting-binding motif.
1,Hemoglobin is a multisubunit allosteric (变构 )
protein that carries O2 in erythrocyte.
1.1 Hemoglobin is a well-studied and well-understood
protein.
1.1.1 It was one of the first proteins to have its
molecular mass accurately determined.
1.1.2 The first protein to be characterized by
ultracentrifuge.
1.1.3 The first protein to be associated with a
specific physiological function.
1.1.4 The first protein with a single amino
acid substitution being related to a genetic
disease (the beginning of molecular medicine).
1.1.5 The first multisubunit protein with
its detailed atomic structure determined by X-
ray crystallography.
1.1.6 The best understood allosteric
protein.
1.2 Determination of the atomic structure of
hemoglobin A (from normal adult) is very revealing.
1.2.1 The protein molecule exists as a a2b2
tetramer,
1.2.2 Each subunit has a structure strikingly
and unexpectedly similar to each other and to that
of myoglobin,indicating quite different amino acid
sequences can specify very similar 3-D structures.
1.2.3 Extensive interactions exist between the
unlike subunits through noncovalent interactions.
1.2.4 Quaternary structure changes markedly
when O2 binds,Crystals of deoxyhemoglobin shatter
(break) when exposed to O2.
1.2.5 O2 binds to the sixth coordination
position of the ferrous iron (as in myoglobin).
1.3 Hemoglobin is a much more intricate and sentient
(sensitive) molecule than is myoglobin.
1.3.1 The oxygen-binding (dissociation) curve of
hemoglobin is sigmoidal and that of myoglobin is
hyperbolic.
1.3.2 Myoglobin has a higher affinity for O2,
evolved for O2 storage.
1.3.3 Hemoglobin releases O2 efficiently at low
oxygen level tissues (thus evolved for O2 delivery),while
myoglobin does not.
1.3.4 Oxygen binding of hemoglobin shows
positive cooperativity,The binding of O2 (the ligand) at
one heme facilitates the binding of O2 at the other hemes
on the same tetramer (vice versa,unloading of oxygen at
one heme facilitates the unloading of oxygen at the
others),(Negative cooperativity refers to a decrease of
activity.)
1.3.5 Increasing concentrations of H+ (with a
decrease of pH) or CO2 lower the O2 affinity of
hemoglobin (H+ and CO2 has no effect on O2 affinity of
myoglobin),This is called Bohr effect,which helps the
release of O2 in the capillaries of actively metabolizing
tissues,
1.3.6 One molecule of 2,3-diphosphoglycerate (BPG)
binds to the central cavity of one tetramer of hemoglobin,
which lowers its O2 affinity.
1.3.7 BPG plays critical roles in the physiological
adaptation to the lower pO2 available at high altitude.
1.3.8 Fetal hemoglobin (HbF) binds BPG less
strongly than does hemoglobin A (adult) and consequently
has a higher oxygen affinity,
Identity
between
Hba and
Hbb,< 50%
Number of
identity AAs
among Mb,
Hba and
Hbb,27
kPa,kilopascal,1 atm = 760 mm Hg = 14.7 psi = 101.3 kPa.
binding sites occupied [L]
? = ------------------------------- = -------------
total binding sites [L] + Kd
? is a measurable quantity in experiment.
[L] is the free ligand concentration.
Hill plot,The ordinate = n log[L] - log Kd
Cooperativity,nH = 1,no; nH > 1,positive;nH < 1,negative.
Bohr’s effect and its molecular mechanism
Release of O2 in
peripheral tissues
Binding of O2 in lungs with
release of H+ and CO2.
1.4 Cooperativity is a particular case of an
allosteric effect
1.4.1 Allosteric effect refers to the
phenomenon in which a molecule (allosteric
effector) bound to one site on a protein causes a
conformational change in the protein such that
the activity of another site on the protein is
altered (increased or decreased).
1.4.2 H+,CO2,and BPG all show an
allosteric effect for the O2 binding process of
hemoglobin.
1.5 Two models have been proposed to explain the
allosteric regulation phenomena
1.5.1 The sequential model (proposed by
Daniel Koshland,Jr.) hypothesizes that the binding
of one ligand to one subunit changes the
conformation of that particular subunit from the T
state (with a low activity) to the R state (with a high
acitvity),a transition that increases the activity of
the other subunits for the ligand,
1.5.2 The sequential model can be analogized
to the tearing process of postage stamps,
1.5.3 The concerted model (proposed by Monod,
Wyman,and Changeux) hypothesizes that symmetry
is conserved in allosteric transitions (all subunits are
in the same conformation) and the binding of each
ligand increases the probability that all subunits in
that molecule are converted to the R-state (with a high
activity),All-or-none model.
1.5.4 The interplay between these different
ligand-binding sites is mediated primarily by changes
in quaternary structure,The contact region between
two subunits can serve as a switch that transmits
conformational changes from one subunit to another.
T,tense state,circle,less active;
R,relaxed state,square,more active
Concerted,all-or-none Sequential
1.5.5 The functional characteristics of an
allosteric protein are regulated by specific
molecules in its environment,In other words,in
the evolutionary transition from myoglobin to
hemoglobin,a macromolecule capable of
perceiving information from its environment has
emerged.
1.6 Sickle-cell anemia was found to be caused by a single
amino acid change in the b chain of hemoglobin molecules.
1.6.1 The hemoglobin molecule from sickle-cell
anemia patients (HbS) was found to have a higher pI value
(having more net positive charges).
1.6.2 Peptide fingerprinting (protease digestion +
electrophoresis + chromatography) of HbS and HbA (wt)
revealed that all but one of the peptide spots matched.
1.6.3 Amino acid sequencing revealed that HbS
contains Val instead of Glu at position 6 of the b chain!
1.6.4 DNA sequencing of the genes further revealed
that the codon of b6 changed from GAA(Glu) to GTA(Val).
1.6.5 The oxygen binding affinity and allosteric
properties of hemoglobin are virtually unaffected by this
change (the b6 is located at the surface of the protein).
1.6.5 High concentration of deoxygenated HbS
forms fiber precipitates,which sickles the red blood
cells,because the fiber formation is a highly concerted
reaction.
1.6.6 Presence of Val6 on the b subunits generates
a hydrophobic patch on the surface which
complements with another hydrophobic patch formed
only in deoxygenated HbS,thus generating the fiber
aggregates (a polymer of HbS).
1.6.7 Sickle cell trait (heterozygote) confers a
small but highly significant degree of protection against
the most lethal form of malaria (probably by
accelerating the destruction of infected erythrocytes,in
Africa).
1.6.8 Fetal DNA can be analyzed for the presence
of the HbS gene (prenatal DNA diagnosis).
1.7 Thalassemias are genetic disorders
characterized by defective synthesis of one or
more hemoglobin chains.
This can be caused by a missing gene,
impaired RNA synthesis or processing,
generation of grossly abnormal proteins.
Mutation and molecular interaction’s changes
2,Immunoglobulin superfamily members,found on cell surfaces
or secreted,are widely used for specific molecular recognition.
2.1.1 Class I MHC proteins bind and display
peptides derived from proteolytic degradation and
turnover of proteins that occurs randomly within the
cell,while Class II MHC detect peptides derived not
from cellular proteins but from external proteins
ingested by the cells.
2.1.2 Class I MHC occurs on the surface of
virtually all vertebrate cells,while Class II MHC is
found on the surfaces of a few specialized cells that
take foreign antigens,including macrophages and B
lymphocytes.
2.1 Major histocompatibility complex (MHC) detects
foreign protein antigens.
2.1.3 Class I MHC has up to 6 variants,while
each human is capable of producing up to 12 variants
of Class II MHC.
2.1.4 The complexes of Class I MHC and its
binding peptide are the targets of the T-cell receptors
of Tc cells in the cellular immune system,while those
for Class II MHC are the binding targets of the T-cell
receptors of Th cells.
2.1.5 Both Tc and Th cells undergo stringent
selection during the maturation process.
2.1.6 T cells patrol the infections foreign antigens,
and have life time of only a few days,Only a minority
of the T cells will trigger full immune response.
2.2 An immunoglobulin G (IgG) molecule (of ~150
kD) was found to contain two light and two heavy
chains,connecting to each other through disulfide
bonds.
2.2.1 Papain digestion convert the IgG
molecule into two Fab (antigen binding) and one Fc
(crystallize) fragments.
2.2.2 Each Fab fragment (containing one
complete light chain and half of a single heavy
chain) binds one molecule of antigen in a similar
manner to the original immunoglobulin molecule.
2.2.3 Each IgG molecule binds to two
molecules of antigens (thus called bivalent).
2.3 Complete amino acid sequence analysis of purified
myeloma patient’s immunoglobulins revealed strikingly
that the L and H chains consist of variable and constant
regions.
2.3.1 Residues 1 to 108 in the L chains are
relatively variable,and 109 to 204 relatively constant.
2.3.2 Residues 1 to 108 in the H chains are
variable,and 109 to 446 relatively constant.
2.3.3 Three segments in the L chain and three in
the H chain display far more variability than do others,
which are thus named as hypervariable segments (also
called complementary-determining regions,or CDRs,
because they determine antibody specificity).
2.3.4 Amino acid sequence analysis revealed that
the variable region of the light chain (VL) is similar in
sequence to that of the heavy chain (VH).
2.3.5 The constant region of each heavy chain
can divided into three parts (CH1,CH2,CH3) of
similar sequences.
2.3.6 The amino acid sequence of the constant
region of each light chain (CL) is similar to that of the
three parts in the constant region of the heavy chains.
Rodney Porter and Gerry Edelman were
awarded the Nobel Prize in 1972 in Medicine or
Physiology for their structure-function studies on the
antibody molecules.
2.4 X-ray crystallography studies revealed,strikingly,
that the immunoglobulin molecules are made of 12
structurally similar domains.
2.4.1 Each domain has a recurring structural
motif,called the immunoglobulin fold consisting of two
broad sheets of antiparallel b-strands joined by a
disulfide bond.
2.4.2 The CDRs of both VL and VH are located
in loops at one end of the sandwich made of the two b-
sheets that come together to form one antigen binding
site.
2.4.3 The immunoglobulin core serves as a
framework that allows almost indefinite variation of
the CDR loops,corresponding to various antigen
specificity of various antibodies.
2.5 T cell receptors,Class I and Class II Major
histocompatibility complex (MHC) proteins,and
intracellular adhesion molecules (ICAMs) all contain
domains similar to the immunoglobulin domains,thus
belonging to the immunoglobulin superfamily.
2.5.1 The immunoglobulin domains (folds) are
widely observed in these protein molecules,revealed by
sequence homology (~20% identity) and similar foldings
in some are confirmed by X-ray structure determination.
2.5.2 All these proteins are involved in molecular
recognition (antigen recognition by antibodies,TCR,and
MHC proteins; cell-cell interactions by ICAMs).
2.5.3 Most of the immunoglobulin superfamily
members are found on cell surfaces (e.g.,IgM,TCR,
MHC) or secreted (IgG).
3,ATP-driven conformational cycles lead to muscle
contraction.
3.1 Striated muscles contain overlapping arrays of
thick and thin filaments.
3.1.1 The thick filaments are primarily made of
myosin protein (肌球蛋白 ).
3.1.2 The thin filaments are primarily made of
actin (肌动蛋白 ),tropomyosin (原肌球蛋白 ),and the
troponin (肌钙蛋白 ) complex.
3.1.3 Thick and thin filaments slide past each
other in muscle contraction (a model proposed based
on X-ray,light microscope,and EM studies,fig.)
3.2 The force of muscle contraction arises from
the interplay of myosin,actin,and ATP.
3.2.1 Myosin consists of two globular
heads joined to a long a-helical coiled coil tail.
3.2.2 The myosin molecule can be cleaved
by trypsin into two partially functional
fragments,with one being able to form
filaments,and the other being ATPase and able
to bind actin.
3.2.3 Myosin molecules spontaneously
assemble into filaments in solutions of
physiological ionic strength and pH.
3.2.4 Actin molecules exist in monomer form (G-
actin) at low ionic strength and polymerize into a
fibrous form (F-actin,very similar to the thin filaments)
at physiological (higher) ionic strength.
3.2.5 Threads of actomyosin (肌动球蛋白 )
complex are formed when mixing actin and myosin in
solution.
3.2.6 Addition of ATP dissociates actomyosin into
actin and myosin.
3.2.7 The actomyosin threads contract when
immersed in a solution containing ATP,K+,and Mg2+,
whereas threads formed from myosin alone do not.
3.2.8 Myosin,being an ATPase,can be
regarded as a mechanoenzyme catalyzing the
conversion of chemical bond energy into
mechanical energy.
3.2.9 The ATPase activity of myosin is
markedly enhanced when it binds to the
polymerized form of actin (F-actin).
3.2.10 The hydrolysis of ATP drives the
cyclic association and dissociation of actin and
myosin,
3.3 The power stroke in contraction is driven by
conformational changes in the myosin head.
3.3.1 In resting muscle,the myosin heads,with
bound ADP and Pi,are unable to interact with the actin
units in thin filaments because of steric interference by
tropomyosin,a regulatory protein.
3.3.2 When muscle is stimulated,tropomyosin
shifts position,and the myosin head (with bound ADP
and Pi) reaches out from the thick filament and interact
with the actin units on thin filaments.
3.3.3 The binding of myosin-ADP-Pi to actin leads
to the release of ADP and Pi,which induces a major
conformational change in the myosin head pulling the
actin filament forward for about 100 Angstroms.
3.3.4 Subsequently,ATP binds to the
myosin head (myosin prefers binding to ATP than
to actin) and thus detaches it from actin.
3.3.5 Finally,the bound ATP is hydrolyzed
by the free myosin head,resetting it for the next
interaction with the thin filament.
3.3.6 The essence of the process is a cyclic
change both in the conformation of the myosin
head and its affinity for actin.
3.3.7 The control of protein-protein
interactions by bound nucleotides (ATP,GTP,etc)
is a recurring theme in biochemistry.
3.4 Troponin and tropomyosin mediate the
regulation of muscle contraction by Ca2+.
3.4.1 Ca2+ is released from sarcoplasmic
reticulum in muscle cells at the stimulation of a
nerve impulse.
3.4.2 Ca2+ controls muscle contraction by
an allosteric mechanism (through
conformational changes) in which the flow of
information is in the following order,Ca2+,the
troponin complex,tropomyosin,actin,myosin,
3.5 Other accessory proteins maintain the
architecture on the myofibril and provide it with
elasticity.
3.5.1 Springlike titin (肌联蛋白 ) molecules
(2.7 MD,or more than 25,000 amino acid
residues),the largest protein so far found in
nature,extend from the thick filaments to the Z
disc.
3.5.2 Nebulin,another large protein
molecule,is closely associated with the actin thin
filaments,and consists of almost entirely of a
repeating 35-amino-acid acting-binding motif.
