1A: Structure & function of proteins and their constituent amino acids Flashcards
Amino Acids
Contain a carboxylic group, alpha carbon, alpha amino group and alpha hydrogen
Absolute Configuration at the α position (Optical Activity)
D (+) = clockwise rotation of polarized light
L (-) = counterclockwise rotation of polarized light
Naturally occurring amino acids
L-Amino Acids
Absolute Configuration at the α position (Stereochemistry)
R (right) vs S (left)
Amino Acids as Dipolar Ions
Low pH = cationic
High pH = anionic
Isoelectric Point = Neutral Zwitterion
Acidic Amino Acids [2] (-)
Aspartic Acid (Aspartate) Glutamic Acid (Glutamate)
Basic Amino Acids [3] (+)
Arginine
Lysine
Histidine
Hydrophobic/Lipophilic Amino Acids [8]
Alanine (A) Valine (V) Leucine (L) Isoleucine (I) Proline (P) Methionine (M) Phenylalanine (F) Tryptophan (W)
Hydrophilic/Lipophobic Amino Acids [12]
Glycine (G) Serine (S) Threonine (T) Arginine (R) Asparagine (N) Aspartate (D) Glutamate (E) Glutamine (Q) Cysteine (C) Lysine (K) Histidine (H) Tyrosine (Y)
Sulfur Linkage Reaction
Cysteine-SH + HS-Cysteine -> Cystine-S-S-Cystine
Importance of cystine
Important for tertiary structure
How are peptide bonds formed?
The carboxyl group of one amino acid reacts with the amino group of a second amino acid; releases water a product
How are peptide bonds broken?
A water molecule is introduced into the peptide bond releasing a free amino acid from the peptide chain
Primary Structure of Proteins
Linear sequence of amino acids, linked by peptide bonds
Secondary Structures of Proteins
Consists of alpha helices and beta sheets; linked by hydrogen bonds
Tertiary Structure of Proteins
Chains of peptides folded onto themselves, linked by disulfide bonds, ionic interactions, van der waals, hydrogen bonds
Importance of Proline
Introduces kinks that cause turns
Importance of Cysteine/Cystine
Forms disulfide bonds
Hydrophobic Bonding
Occurs within the core of proteins between the non-polar/hydrophobic R-groups creating stability (hydrophobic collapse)
Quaternary Structure of Proteins
3D structure with multiple subunits of proteins interacting; linked by non-covalent interactions between subunits
Conformational Stability
The dG difference between the native state (folded) and unfolded state of a protein
Denaturation
Occurs due to temperature, chemicals, enzymes and pH
How does temperature denature?
It disrupts all bonding expect peptide bonds; this increases hydrophobic interactions since active globular proteins will fold
How do chemicals denature?
They break hydrogen bonds, disrupts all except peptide bonds
How do enzymes denature?
They break down directly to peptide bonds
How does pH denature?
Ionic bonds are broken down so tertiary and quaternary structures are disrupted
How does a solvation layer affect stability?
It decreases the amount of ionic interactions between proteins
Isoelectric Point (Separation)
Proteins move until they reach the pH equal to their isoelectric point in electrophoresis
Electrophoresis (Separation)
Separates charged particles using an electric field
Agarose Gel Electrophoresis
Separates nucleic acids; their negatively charged structures move toward the cathode and help with identification of sizes of particles
SDS-PAGE
Separates proteins based on mass but not charge; SDS neutralizes charge; smaller particles move through the gel faster
Non-Enzymatic Protein Function
Binding of molecules
Immune Function (Ab)
Movement (Dynein & Kinesin)
Transport (Hemoglobin)
Function of Enzymes in Biological Reactions
They act as catalysts, providing alternate pathways for reactions to occur; stabilize transition states
Types of Catalysis
Acid/Base
Covalent
Electrostatic
Acid/Base Catalysis
Acids donate protons, bases accept protons
Covalent Catalysis
Formation of covalent bonds in order to reduce energy for transition states; bonds are broken for the reuse of the enzyme
Electrostatic Catalysis
Formation of ionic bonds with intermediates in order to stabilize the transition states in chemical reactions
Types of Enzymes
- Oxidoreductases
- Transferases
- Hydrolases
- Isomerases
- Lyases
- Ligases (Synthetases)
Oxidoreductases
Transfers hydrogen and oxygen atoms or electrons from one substrate to another
e.g. Dehydrogenase, Oxidase
Transferases
Transfer of a specific group from one substrate to another
e.g. Transaminase, Kinase
Hydrolases
Hydrolysis of a substrate
e.g. Esterases, Digestive Enzymes
Isomerases
Change of the molecular form of the substrate
e.g. Phosphoglucoisomerase, Hexoisomerase, Fumarase
Lyases
Nonhydrolytic removal of a group or addition of a group to a substrate
e.g. Decarboxylase, Aldolase
Ligases (Synthetases)
Joining of 2 molecules by the formation of new bonds
e.g. Citric Acid Synthetase
Reduction of Activation Energy
Enzymes reduce the energy of activation by providing alternate pathways for reactions; which increases the rate of the reaction
Saturation Kinetics
The idea that as concentration of substrate increases, so does the rate of the reaction
What do enzymes NOT affect?
Keq, dG & Thermodynamics
What do enzymes affect?
Rate Constant, Kinetics, Forward & Reverse reaction (no change in equilibrium)
Substrate Specificity
Substrate binds at the enzymes active site; their structure is specific to fit into the enzymes active site
Active Site Model of Enzyme Specificity
The enzymes active site has a shape that accommodates the shape of the substrate
Induced-Fit Model of Enzyme Specificity
Enzymes and their substrates conform to each others’ shape in order to bind together
Cofactors
Inorganic molecules or Metal ions that certain enzymes use to catalyze a reaction/process
Holoenzyme
Enzyme + cofactor
Apoenzyme
Enzyme - cofactor
Prosthetic Group
Tightly bound coenzyme
Cosubstrates
Loosely bound coenzyme
Coenzyme
Small, organic, non-protein molecules that carry chemical groups (electrons, atoms, functional groups) between enzymes; vitamin derivatives
Water-Soluble Vitamins
B Complex (B1, B2, B6, Folate, B12, Biotin, Pantothenate) C
Fat-Soluble Vitamins
Vitamin A, D, E, K
How do local conditions affect enzyme activity?
pH, salt, temperature etc, can all affect the structure of the enzymatic protein and thus affect the availability of the active site
Michaelis-Menten Kinetics Equation
E + S -> ES -> E + P
Michaelis-Menten Approximations
Rapid Equilibrium & Steady-State
Rapid Equilibrium Approximation
It states that E, S and the ES complex equilibrate rapidly so that the total enzyme concentration is equal to the concentration of free enzyme and the concentration of bound enzyme;
Etotal = Efree + ES
Steady State Approximation
That the rate of formation of the ES complex is equal to the rate of breakdown of the ES complex
Km (Michaelis Constant)
Breakdown[ES]/Formation[ES]
Factors that affect Km
pH, temperature, ionic strength, nature of substrate
Reaction Order (Enzyme Kinetics)
Zero Order; Rate is independent of substration formation
Vmax/2 (1/2 Vmax)
Km
Units of V
Moles/Time
Units of Substrate
Molar
Low Km indicates:
Not much substrate required to reach half maximal velocity; high affinity for the particular substrate
High Km indicates:
A lot of substrate required to reach half maximal velocity; low affinity for the particular substrate
Cooperativity
When a substrate binds to one subunit, the other subunits are stimulated and become active. It can be positive or negative
Positive Cooperativity & it’s Curve
One oxygen molecule binds to the ferrous iron of a heme molecule which allows the other subunits heme to bind more molecules; Sigmoidal Shape
Feedback Regulation
The product of a pathway inhibits or activates its pathway; it can be positive (activation) or negative (inhibition)
Competitive Inhibition
Inhibitor competes with its substrate for the active site; can be overcome by increasing the amount of substrate; the Vmax is unchanged by the inhibitor; apparent Km increases
Noncompetitive Inhibition
Inhibitor binds to the enzyme at an allosteric site which deactivates it; substrate still has access to the A/S but cannot catalyze the reaction as long as the inhibitor binds;
Decreases Vmax; Unchanged Km
Mixed Inhibition
Inhibitor can bind to the allosteric site or the ES complex;
Decreases Vmax; Increases or Decreases Km
Uncompetitive Inhibition
Inhibitor binds only to substrate-enzyme complex; Decrease Vmax; Decreases Km
Allosteric Enzymes
Contain 2 binding sites, one for substrate & others for effectors (which change the conformation of the enzyme, noncovalently & reversibly)
Homotropic Allosteric Enzymes
Acts as both the substrate for the enzyme and the effector of the enzyme’s activity
Heterotropic Allosteric Enzymes
Acts only as the effector that regulates the enzyme’s activity; does not act as substrate
Covalently-Modified Enzymes
Covalent modification (phosphorylation) activates or inactivates the enzymes activity e.g. Glycogen phosphorylase-a vs b
Zymogen
Inactive enzyme precursor; upon hydrolysis or change of configuration of the active site
