Transcript Chapter 3 Proteins:

Chapter 3 Proteins:
Shape, Structure, and Function
Proteins Execute Cell Functions
Enzymes
► Channels and pumps
► Signal Molecules
► Messengers
► Molecular Machines
► Structural Support
► Cell Recognition
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Protein Shape and Structure
Peptide Bond Links Amino Acids into Polypeptide Chain
Protein Shape and Structure
Evolution fine-tuned structure and chemistry
► Shape dictated by amino acid sequence
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polypeptide backbone
side chains
Protein Shape and Structure
Sequence Determines Structure
Protein Shape and Structure
Weak Noncovalent Bonds/Interactions
important to the folding of polypeptide chain
Protein Shape and Structure
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Fold into Conformation of Lowest Energy
Common Folding Patterns
alpha helix
Beta Sheet
Coiled Coils
Protein Shape and Structure
Levels of organization protein structure
primary= aa seqeunce
► secondary= stretches of alpha helix, beta sheets
► tertiary=3d organization
► quartenary=complete structure of protein w/ > 1 polypeptide chain
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Protein Shape and Structure
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Protein Domain= Fundamental Unit of Organization
independently folding unit
40-350 aa modular unit; combine to form larger proteins
different domains have different functions
Fold= central core of domain; comprised of beta sheets and alpha
helices in various combinations; limited number
Short signature sequences identify homologous protein domains
Protein Shape and Structure
Domain shuffling during the course
of evolution
Percentage of total genes in
respective genomes containing
one or more copies of a
particular protein domain
Protein Shape and Structure
Protein Module
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Smaller than an average domain, generally 40-200 aa
Particular versatile structures
Easily integrated into other proteins; form parts of many
different proteins
Protein Shape and Structure
Protein Families Evolved
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similar 3d structure
portions or aa sequence conserved
non-conserved portions impart new functionality
serine proteases
homeodomain proteins
kinases
immunoglobulins
Protein Shape and Structure
► Sequence
Homology Searches
► Amino Acids Sequence Threading
► Modules form parts of many different proteins
Protein Shape and Structure
Protein Shape and Structure
Larger proteins can assemble from identical monomeric subunits
Protein Shape and Structure
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Larger proteins often contain more than one polypeptide
Proteins can serve as subunits for assembly of large
structures
Self Assembly
Protein Function
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Function of protein dictated by physical interactions w/
other molecules
specificity and ligand affinity governed by multiple weak
noncovalent bonds
active/binding site often cavity on protein surface formed by neighboring
aa or aa that may belong to different portions of polypeptide
Protein Function
Conformation determines chemistry
Regions adjacent to active or ligand binding site may restrict water to increase ligand
binding
► Clustering of polar or chged residues can alter chemical reactivity
► Type and orientation of exposed aa side chains govern chemical reactivity
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Protein Function
“Evolutionary tracing” to determine sites critical to protein function
3d structure of protein family members are similar even when aa homology
falls to 25%
► Map unchg aa or nearly unchg from all known family members onto 3d
structure of one family member
► Most invariant positions often on surface and represent ligand binding site
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Protein Function
Proteins Bind to other Protein Through Several Types of Interfaces
Protein Function
Equilibrium Constant Describes Binding Strength
Steady state or equilibrium:
# association events/sec = # dissociation/sec
► From conc of two molecules and complex equilibrium constant can be calculated
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Protein Function
Enzymes as Catalysts
Make or break covalent bonds
► Speed up chemical reactions > 106 fold
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Stabilize transition state
Decrease activation energy
Increase local conc of substrate at catalytic site
Hold reactants in proper orientation for chem rxn
Binding energy contributes directly to catalysis
Not consumed or changed during process
Protein Function
Common Types of Enzymes
Hydrolases
Nuclease
Proteases
Phosphatases
Isomerases
Polymerases
Kinases
OxidoReductases
ATPases
Synthases
Protein Function
Enzyme Kinetics
Vmax= how fast enzyme can process substrate, pt at which enzyme saturated w/substrate
► Turnover Number= Vmax/[enzyme]
turnover ranges from 1-10,000 substrate molec/sec
► Km= substrate conc at Vmax/2; measure of affinity
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Protein Function
Lysozyme
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Natural antibiotic in egg white, tears, saliva
Hydrolyzes polysaccharide chains residing in cell wall of bacteria
Protein Function
Specific Mechanism of Lysozyme Hydrolysis
Enzyme positions substrate bending critical chem bonds that participate in chem rxn
► Positions acidic side chain of Glu w/in active site to provide high conc of acidifying H+ ions
► Negatively chged Asp stabilizes positive chged transition state
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Protein Function
General Mechanism for Enzyme Activity
Active site contains atoms that speed up rxn
► Substrate driven towards transition state upon binding to enzyme; shape of
substrate chgs & critical bonds bent
► Covalent bond sometimes formed btwn substrate and side chain of enzyme
► Restoration of side chain to original state
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Protein Function
Small Molecules Add Extra Functions to Proteins
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Chromophores detect light; retinal
Metal atoms assist w/ catalytic functions; Zn, Mg, Fe
Coenzymes (sm organic molec) provide functional grps; biotin
Protein Function
Multienzyme Complexes
Increase the rate of cell metabolism
► Product of enzyme A passed directly to enzyme B; product of enzyme B passed to
enzyme C; and so on
► Simulates intracellular membrane compartment; effectively increasing substrate conc at
site of enzyme activity
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Protein Function
Regulation of Catalytic Activity
Negative Feedback
► Positive Regulation
► Allosterism
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Protein Function
Allosterism
Protein Function
Symmetric Protein Assemblies and Cooperative Allosterism:
sm chgs in ligand conc switches enzyme assembly from fully active to fully inactive state via
conformation changes that are transmitted across neighboring subunits
Protein Function
Allosteric Transition in Aspartate Transcarbamoylase
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6 catalytic subunits and 6 regulatory subunits
all or none transition between T-tense and R-relaxed state
Active R state driven by binding of carbamoylphosphate and aspartate
Inactive T state driven by binding of CTP to regulatory dimers
Protein Function
Regulation by Phosphorylation/Dephosphorylation
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Addition or removal of P grp carrying (2) negative chgs can
cause major conformation chg in protein
Phosphorylation/dephosphorylation of proteins= response to
signals that specify chg in cell state
Protein Function
Protein Kinase
transfers terminal P of ATP to OH grp of SER, Thr, or Tyr
► 100’s ea specific for particular target
► Kinases share 250 aa catalytic domain
► Non-conserved aa flanking catalytic site or in loops w/in kinase domain confer specificity
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Protein Function
Protein Phosphatases
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Catalyzes the removal of P grp
Some specific; some act on broad range of proteins
Protein Function
Protein can Function as Microchip
Cdk= cyclin dependent protein kinase activity
dependent upon 3 events:
1. binding of second protein cyclin
2. phosphorylation of Thr side chain
3. dephosphorylation of Tyr side chain
Cdk monitors specific set of cell components
acting as input-output device
Protein Function
GTP Binding Proteins
Analogous to Proteins regulated by P/de-P
► Active when GTP bound; inactive when GTP hydrolyzed
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Protein Function
Regulatory Proteins Control Activity of GTP Binding Proteins
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GAP= GTPase activating protein; binds and induces hydrolysis
GEF= Guanine nucleotide exchange factor; binds to GDP protein causing it to release GDP in
exchange for GAP
Protein Function
Large Protein Movements Generated from Small Ones
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EF-Tu
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= elongation factor in protein synthesis, GTPase
tRNA complexes w/ GTP bound form of EF-Tu w/ aa masked
GTP hydrolysis occurs when tRNA binds to mRNA on ribosome; tRNA disassociates
GTP hydrolysis causes “Swtich helix” to swivel unmasking aa
Protein Function
Motor Proteins
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Produce lg movements in cells such as:
muscle contraction
crawling and swimming of cells
movement of chromosomes
movement of organelles
enzymes on DNA
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Possess moving parts as force generating machines
Protein Function
ATP hydrolysis allows unidirectional series of conformational chgs to propel proteins along DNA
Protein Function
Allosteric proteins harness energy derived from ATP hydrolysis, ion
gradients, electron transport processes to pump ions or sm molecules
across membranes
Ca2+ Pump of Sarcoplasmic Reticulum
Protein Function
Mechanism of Ca2+ Pump
Protein Function
Structure of Ca2+ Pump
10 transmembrane helices
► 4 transmembrane helices provide
Ca2+ binding sites for pump
► helices that bind Ca2+ wind around
ea other forming cavity btwn helices
for Ca2+
► ATP hydrolysis causes conformation
chgs that later cavity enabling Ca2+
to be pushed through
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