Exam 2 Ch.4: Protein Structure and Function Flashcards
(101 cards)
1
Q
Enzymes
A
catalyze covalent bond breakage or formation
2
Q
Structural Proteins
A
provide mechanical support to cells and tissues
3
Q
Transport Proteins
A
carry small molecules or ions and transport them throughout the body
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Q
Motor Proteins
A
generate movement in cells and tissues
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Q
Storage Proteins
A
store amino acids or ions
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Q
Signal Proteins
A
carry extracellular signals from cell to cell
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Q
Receptor Proteins
A
detect signals and transmit them to the cells response machinery
8
Q
Transcription Regulators
A
bind to DNA to switch genes on or off
9
Q
Special-Purpose Proteins
A
highly variable
10
Q
Disulfide bond
A
can form between two cysteine side chains
11
Q
Primary Level of Protein Structure
A
Chain of amino acids
12
Q
What dictates the overall 3D shape of a protein?
A
side chains
13
Q
What stabilizes protein conformation?
A
- the sum of weak forces
-van der Waals, electrostatic attractions, hydrogen bonds,
14
Q
Backbone to backbone bond
A
hydrogen bond
15
Q
What governs specific folding of proteins?
A
- making the most energetically favorable bonds
- meaning proteins folds into the lowest free energy conformation
- folding is a spontaneous process and will release heat
16
Q
Example of Spontaneous Protein Folding and Refolding
A
- Protein is exposed to high concentration of urea
- causes denatured protein
- remove urea
- protein refolds
17
Q
Chaperone Proteins
A
- assist with protein folding
- spontaneous folding can be time-consuming so chaperones speed up protein folding rates
- come bind to partly fold chains
18
Q
Benefit and Drawback of Isolation Chambers
A
- can help prevent aggregation of multiple proteins
- can isolate a single protein
- takes more energy
19
Q
How do isolation chambers work?
A
- one polypeptide chain is sequestered by the chaperone
- chamber cap closes chaperone
- isolated polypeptide chain folds correctly
- correctly folded protein is released when cap dissociates
20
Q
What stage is a protein done in the chaperone protein?
A
depends because proteins show incredible structural diversity
21
Q
What are some basic functions of proteins?
A
transport, structure, signaling, catalyzing reactions
22
Q
For a polypeptide chain:
What is the structure of its backbone?
Which end is the 1st amino acid? The last?
A
- they have an N-C-C backbone
- 1st amino acid is the N-terminus
- last amino acid is the C-terminus
23
Q
What are the forces that govern a strict and specific folding pattern of a single protein?
A
- proteins folding into the lowest free energy conformation to make the most energetically favorable bonds
24
Q
What bonds are responsible for protein folding? Where can these bonds occur?
A
- weak forces such as electrostatic attractions, hydrogen bonds, and van der Waals are responsible for protein folding
- they can occur backbone to backbone, backbone to side chain, side chain to side chain
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Secondary Protein Structure
Alpha helix and beta sheet
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What stabilizes alpha helices?
- stabilized by hydrogen bonding along the backbone
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Structure of Alpha helices?
- amino and carboxyl groups of every 4th amino acid engage in hydrogen bonding
- r groups pointed outwards
- can be right or left handed
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bonds from backbone to backbone
hydrogen bond between atoms of two peptide bonds
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backbone to side chain
hydrogen bond between atoms of a peptide bond and an amino acid side chain
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side chain to side chain
hydrogen bond between atoms of two amino acid side chains
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Tertiary Protein Structure
- 3D arrangement of secondary structures
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Why is it good that the R groups in alpha helices point outward?
- Because alpha helices are commonly found in membranes
- Hydrophobic R-groups point outward away from the core of the helix and towards hydrophobic membrane
- backbone is typically hydrophilic
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What forms coiled- coils and why are they beneficial?
-2-7 alpha helices can form coiled- coils
- helices wrap around each other to minimizes exposure of hydrophobic amino acids side chains to aqueous environment
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What holds together beta sheets?
- held together by hydrogen bonds between the strands (backbone of strand)
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Direction of beta sheets and r groups?
R-groups poke out on alternating sides (pleated) (top-bottom-top-bottom...)
-can be parallel or antiparallel
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Consequence of misfolding proteins?
- can lead to aberrant protein structures which build up and lead to cell death
- causes diseases like Alzheimers, Parkinson's, Huntingtons
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Example of misfolding causing disease?
In Alzheimers and Parkinsons disease, insoluble amyloid fibers are made from misfiled alpha-synuclein, which builds up to a toxic concentration and kills neurons
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What are prions?
an infectious form of misfodled proteins
- Ex: mad-cow disease, chronic wasting disease, Creutzfeldt-Jakob disease
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How do prions work?
can induce normal proteins of the same type to misfiled in to the same misfolded form
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What would the differences in R-groups be between an alpha-helix in a cytoplasmic protein versus an alpha-helix in a transmembrane protein?
cytoplasmic protein: alpha helices in a coiled coil to minimize exposure of hydrophobic amino acid side chains to aqueous environment
transmembrane protein: hydrophobic r-groups point outward away from the core of the helix, backbone is usually hydrophilic
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What forces form a coiled-coil?
2-7 Alpha helices that wrap around each other to minimize exposure of hydrophobic amino acids side chains to aqueous environment
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What makes a Beta-sheet pleated? Parallel or anti-parallel?
pleated - r-groups poke out of alternating sides
Parallel vs antiparallel: n-terminus/ c-terminus going the same way vs not
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Tertiary Structure
3D arrangement of secondary structure
Main types: fibrous and globular
- Globular: enzymes, etc
- Fibrous: structural/ strength
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What is a protein domain?
any segment of a polypeptide chain that can fold independently into a compact stable structure
- associated with specific functions (DNA binding domain, catalytic activity, etc.
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Effect of mutation in a domain?
mutation in a specific domain can affect the function of that domain
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Protein Families
Families have proteins with similar structures, but over time have gained slight differences that give them slightly different functions
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Quaternary Structure
- multiple subunits (optional)
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Subunit
individual folded polypeptide chain
- dimer/trimer/tetratrimer
- Heterodimer vx homodimer
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Homodimer
two proteins with the same amino acid sequence that bind together to form a function
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Heterotetramer
- four individual subunit with differing amino acid sequences that work together to perform a function
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What are the types of structures that subunits form?
- helices, rings
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Actin fibers
helical structures made up of many individual actin proteins
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Microtubules
long hollow tubes of the protein tubulin
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Protein Cross-Linking
- what it does
- where it's common
- example
- proteins can be stabilized by covalent cross linking
- very common in extracellular matrix proteins
- a type of cross linking is the formation of disulfide bond between cysteine amino acids
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Elastin
an ECM protein that's stabilized by covalent cross links
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How do proteins work?
their properties/functions depend on physical interactions with other molecules
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Ligands
a substance that forms a complex with (binds to) a specific protein
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How do ligands form a complex?
-forms a complex thanks to the sum of its non-covalent interactions
- these interactions define is specificity, "Hand in glove" or "Lock in Key"
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Structure of IgG antibodies?
are made up of 2 heavy chains, and 2 light chains
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Antigen
- something that causes an immune response
(each antibody can recognize one specific antigen)
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Antigen binding sites
-where antigens bind to an antibody
- antigens are bound at the antigen-binding sites at two ends of an IgG antibody
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Antibody specificity
each antibody binds with a distinct antigen
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Structure of an antibody?
- 2 heavy chain, 2 light chain
- constant domains: sequences constant from antibody to antibody
- variable domains: vary between antibodies; antigen binding domains vary between antibodies
- structure determines function
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How do antibodies defend us against infection?
- antibodies cross-link antigens into aggregates (clump together)
- antibody-antigen aggregates are ingested by phagocytic cells
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Uses of Antibodies in research?
1) molecule purification
2) protein visualization
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Molecule Purification
- using antibodies to purify molecules
- immunoprecipitation
- immunoaffinity column chromatography
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Protein visualization
once antibodies are bound to protein of interest with fluorescent tag, we can shine specific light and see the protein
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Steps of molecule purification? Know both types
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What are 3 ways that enzymes can catalyze a reactions?
(a) enzyme binds to two substrate molecules and orients them precisely to encourage a reaction to occur between them
(b) binding of substrate to enzyme rearranges electrons in the substrate, creating partial negative and positive charges that favor a reaction
(c) enzyme strains the bound substrate molecule, forcing it towards a transition state that favors a reaction
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Vmax
maximum rate of a reaction
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KM
the concentration of a substrate where an enzyme works at half its maximum speed
- small KM: strong binding (strong affinity)
- large KM: weak binding
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Enzyme reaction rate
determined by substrate concentration and affinity
- measured by Vmax and KM
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Therapeutics
- drugs mostly target enzymes
ex: Rifampin and Gleevac (Imatinib)
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Coenzymes
small molecules that aid the function of enzymes
- ex: heme groups bound to hemoglobin (heme groups reversibly bind to oxygen gas through its iron atom) (without heme, hemoglobin can bind oxygen)
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Why is it important to regulate protein activity?
- prevents waste
- some reactions are only beneficial at certain times and in certain cellular locations
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Proteasome Parts
- regulatory cap (blocks core to stop proteasome from destroying stuff)
- catalytic core
- regulatory cap
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1st Strategy to regulate protein activity
- balance proteins production (transcription/translation) and degradation
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2nd Strategy to regulate protein activity
Control the location of the protein
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3rd Strategy to regulate protein activity
Feedback inhibition
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4th Strategy to regulate protein activity
Phosphorylation of the protein
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Process of Feedback Inhibition
- the end product of a downstream reaction binds and inhibits an enzyme earlier in the pathway of the downstream reaction
- binds to a regulatory site on the enzyme
- this is a form of negative regulation (positive regulation also exist)
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How does Feedback Inhibition work?
- works through an allosteric site
- enzymes have at least 2 sites: active site, allosteric site
- binding of inhibitor to allosteric site induces conformational change
- lowers affinity of active site for substrate
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What are the advantages of feedback inhibition?
- Ex. synthesis of amino acids form aspartate
- very specific
- nearly instantaneous
- can be reversed as soon as concentrations of product decrease
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Phosphorylation
- serine, threonine, and tyrosine amino acids side chains can become phosphorylated
- Phosphorylation can activate or inactivate a protein
- causes conformational change
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Kinases vs Phosphatases
- phosphorylation is reversible
- Kinsases: phosphorylation
- Phosphatases: removes phosphate
- phosphate is usually transferred from ATP
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How do protein modifications impact protein function?
Things impacted: activity, stability, binding partners and cellular location
- many proteins have multiple/many sites of covalent modification
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Phosphorylation
on/off switch; also creates docking site for binding partners
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homogenization
gently mechanical procedures where the plasma membrane of cells can be ruptured so that the cell contents are released
Types:
- break apart cells with high-frequency sound
- use a mild detergent to make holes in plasma membrane
- force cells through small hole using pressure
- shear cells between a close-fitting rotating plunger and the thick walls of a glass vessel
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homogenate or extract
thick soup, contains large and small molecules form the cysts, such as enzymes, ribosomes, and metabolites, as well as all of the membrane-enclosed organelle
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Differential Centrifugation
- repeated centrifugation at progressively higher speeds with fractionate cell homogenates into their components
- centrifugation separates cell competes on the basis of size and density. the larger and denser components experience the greatest centrifugal forces and move most rapidly. they sediment to form a pellet at the bottom of the tube, while smaller, less dense components remaining suspension above, a portion called the supernatant
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Gel electrophoresis
when an electric field is applied to a solution containing protein molecules, the proteins will migrate in a direction and at a speed that reflects their size and net charge.
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SDS polyacrylamide-gel electrophoresis (SDS-PAGE)
individual polypeptide chains forma complex with negatively charged molecules of sodium dodecyl sulfate (SDS) and therefore migrate as
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Gel Electrophoresis - Western Blotting
- proteins from gel electrophoresis are transferred onto a membrane
- Antibodies are used to detect a specific protein of interest
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Immunoprecipitation
- mixture of antibodies
- add specific anti-A antibodies
- collect aggregate of A molecules and anti-A antibodies by centrifugation
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Immunoaffinity Column Chromatography
- bead coated with anti-A antibodies
- column packed with these beads
- mixture of molecules flows through column
- flow through is discarded
- elute antigen A from beads
- collect pure antigen a
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Acetylation
on lysine amino acid
- impacts protein: DNA interaction of histones
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Ubiquitination
targets proteins for degradation
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How can antibodies be used in research?
- Molecule Purification
->Immunoprecipitation
->Immunoaffinity column chromatography
- Protein Visualization
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What is Molecule Purification and what types are there?
- uses antibodies to purify molecules
- Immunoprecipitation
- Protein Visualization
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What is Immunoprecipitation
1) mixture of molecules,
2) add specific anti-A antibodies
3) collect aggregate of A molecules
and anti-A molecules by centrifugation
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What is immunoaffinity column chromatography?
1) mixture of molecule
2) goes into column packed with beads coated with anti-A antibodies
3) discard flow through
4) elute antigen A from beads
5) collect pure antigen A
