Week 4-6 Flashcards
Lipids Amino Acids Proteins Enzyme
LIPIDS
- Triglycerides
- Phospholipids: Cholestrol, Liposomes
Lipids:
- Are defined by solubility in non-polar solvents (physical property) rather than by chemical structure
- Many have hydrocarbon/modified hydrocarbon structure, properties + behaviour
Triglycerides/Triesters/Triacylglyceroids:
- Have a glycerol head and 3 fatty acid tails
- Provide energy (which comes from hydrocarbons) and membranes (phospholipids)
- Saturated tails = tightly packed, solid at room temp
- Unsaturated tails = kink, liquid at room temp (plants)
To make unsaturated saturated, undergo hydrogenation
Phospholipids:
- Have 2 fatty acid tails + phosphate group head (-ve charged)
- Therefore one side polar, other side non-polar
- Second most abundant occurring lipid
- Found only in plant+animal membranes (40-50%)
- Mostly derived from phosphaditic acid
- Present in cell membrane
- Regulate what comes in/out eg. cholesterol maintain fluidity of membrane
Cholesterol:
- Allow CO2 out, O2 in by moving left and right
- Cholesterol grabs onto fatty acid tails and hold them together when it is too warm, apart when it’s too cold
- If cell gets heated, phospholipid fall apart
- If cell gets cold, phospholipids crowd together
Liposomes:
- Are formed when phospholipids are shaken with H2O vigorously
- Are small spherical vesicles within a lipid bilayer surrounding an aqueous center
- Water soluble is trapped in centre of liposomes, lipid soluble is incorporated into lipid bilayer
- Useful for drug delivery as it can fuse with cell membranes and empty contents
LIPIDS
- Saponification
- Steroids
Saponification:
- Natural soaps = boiling animal fat with NaOH
- Ester hydrolysis reaction
- Soap molecules are driven by hydrophillic and hydrophobic forces, spontaneously self assemble into micelles, water can’t get in, lipophyllic
Steroids:
- Group of plant+animal lipids that have tetracyclic ring structure
- Are nearly flate
- Quite rigid
- Many hormones = tetrocyclic
Amino Acids
- Structure
- Zwitterion
Amino Acid: General Structure: NH2 - CR - COOH - Central carbon = alpha C - Contain acid and basic group - All naturally occurring = L form (NH2 on left side of alpha C) - Have 8 essential - 19 different R groups
Zwitterion:
- Amino acid undergo acid base reaction to form zwitterion (neutral ion)
- This gives amino acids many physical properties of salts
- In acidic solution, accept H, become basic
- In basic solution, give H, becomes acidic
Amino Acids
- Isoelectric Point (pI)
- Electrophorosis
Isoelectric Point (pI):
- Describes the pH where sample of amino acid where it is neutral
- Is different for each amino acid due to side chain
- If amino acid is submerged in water solution where pH = pI, amino acid –> zwitterion
pI = 1/2 (pKa COOH + pKa NH3)
Electrophorosis:
- Technique for separating charged particles, including amino acids from the protein
- Amino acids migrate towards electrode with charge opposite its own when electric potential is applied
- Higher charged density move faster
- Molecules at pI don’t move
- Strip of paper dried to make amino acids visible
Amino Acids
- Peptides
- Proteins
Peptides:
- Short polymer of amino acids joined by peptide bonds (CONH)
- Classified by number of amino acids in chain
eg. dipeptide, tripeptide, polypeptide - Have resonance structures
- X-Y and Y-X are different
Peptides and proteins are written with
- N terminal: amino terminal amino acid: NH3 on left
- C terminal: amino terminal amino acid: COOH on right
Proteins:
- Biological macromolecule
- Are polymers of amino acids
- Made of 1 or more folded polypeptide chain
Amino Acids
- Proteins:
Primary
Secondary: Alpha helix, Beta sheet, random coil
Tertiary: Hydrophobic interactions, Disulfide bond
Side not: London dispersion force is a temporary attractive force due to temporary dipoles. Weakest intermolecular force
Primary: (peptide bonds, covalent)
- Sequence of amino acids in a polypeptide chain
- Backbone = alternating peptide bonds and alpha C atoms
Secondary: (H-bonds)
- Regular and repeating spatial organisation of neighboring segments of single protein chains
- disulfide bonds ensure protein stays in this structure
- local folded structures that form within a polypeptide due to the interactions between atoms of the backbone
- most common = alpha helix and beta sheet
alpha helix:
- Carbonyl group face down, Nitrogen up, form H -bonds
- 3.6 turns per helix
beta sheet: - antiparallel (strands point in different directions): perfect fit (N terminus is positioned next to the C terminus of the other) - parallel (strands point in same direction): diagonal links (N and C terminus math up)
random coil:
- shorter amino acids allow sharp turn
Tertiary: (Van der Waals, disulfide, London, hydrophobic)
- Overall structure of protein molecule
- Formed by secondary structure folding and bending of chain
- Primarily due to interactions between R group
- Bonds formed include the Van Der Waals and more importantly, hydrophobic interactions and disulfide
- Hydrophobic: amino acids with hydrophobic (nonpolar) R groups cluster together on the inside of protein, leaving hydrophillic amino acids on outside to interact with surrounding H2O molecules (most energy free state)
Quaternary (Van der Waals, disulfide, London, hydrophobic)
- Overall structure composed of more than one polypeptide chain (tertiary structure)
PROTEINS
- Myglobin
- Hemoglobin: Cooperativity
Myglobin:
- Tightly folded protein (tertiary structure)
- 70% helices
- Has iron protoporphyrin group (organic compound) that binds O2 and supply it to the mitochondria of cells
- Hence has high affinity with O2, as it needs to bind tightly to it
- Found in muscle tissue
- Held by non-polar groups, hydrophobic interactions
Hemoglobin:
- Made up of 4 subunits = tetramer
- Has 2 alpha helix and 2 beta sheets, each has a binding site to the heme group (prosthetic group)
(2 subunits = dimer
3 subunits = trimer
many subunits = disomer)
- Similar structure and pattern to myglobin
- Has a quaternary structure to carry O2
- Needs to bind O2 in lungs and release it later, whereas myglobin binds too well
- T state (tense) = Low affinity with O2
- R state (relax) = High affinity with O2
Cooperativity in Hemoglobin:
- Binding of O2 to heme group of 1 hemoglobin subunit causes changes in the conformation of the heme group and surrounding amino acids
(T –> R state in one subunit)
- Since some of the amino acids are in contact with other subunits of protein, this causes change to all subunits
(R state for all subunits)
- This changes all other subuits to increase their affinity for O2
(Hence, R state allows for higher affinity for O2)
- This is positive cooperativity (allosteric behaviour)
Side not: negative cooperativity is the opposite: affinity decreases when the state in all subunits are changed
ENZYME CATALYSIS
- Basic Features & Function
- Reaction Conditions
- Effectiveness
Basic Features & Function:
- Are often proteins with characteristic 1,2,3,4, structure
- Facilitate chemical reactions under biological relevant conditions (eg. pH and temp)
- Increase rate of chemical reaction, and may be recovered unchanged after the reaction is complete
- Catalyse specific reaction (enzyme specificity)
- Role is relevant to 3D shape
Reaction Conditions:
- Low temp (37 in humans)
- Low reactant concentration (10^-3 to 10^-6 M)
- Neutral pH (except stomach: lysosome)
Effectiveness as catalysts:
Ratio = Rate of enzyme catalysed reaction
___________________________
Rate of non enzyme catalysed reaction
= 10^5 to 10^20
ENZYME CATALYSIS (OTHLyILi) - 6 Major Classes Oxidoreductase Transferase Hydrolase Lyase Isomerase Ligase
Oxidoreductases:
Transfer of electrons
(reductase = giving e, oxidase = taking e)
Transferases:
Group transfer reactions
A + BX –> AX + B
Hydrolases:
Hydrolysis reactions, use water to separate A into B + C
(transfer of functional groups to H2O)
Lyases:
Catalyse dissociation of A into B+C without using redox (like oxidoreductants) or water (hydrolases)
eg. urea cycle
Isomerases:
Transforming a molecule to its other isomer
Transfer of groups within molecules to yield isomeric forms
Ligase:
Combine two molecules to form a complex of the two
Ligate = join
ENZYME CATALYSIS
- Active Site
- Binding Site :
Induced fit, Lock and Key
- Catalysis Reactions
NOTE:
Substrate specificity:
Enzyme accept certain substrates
Reaction specificity:
Enzyme undergo specific class of reactions
Active Site:
- specific area of enzyme structure where the substrate binds and chemical groups of the enzyme facilitate the catalysis of the reaction from substrate to products
A + B –> AB –> [X] –> C.D –> C + D
- Old chemical bonds must break
- New bonds must form
- Transition state = neither original/ final molecules are present
- Energy is required to break old bonds to form transition state
Binding Site:
- Must be accommodating to substrate
- Have a complementary 3D shape + chemistry for specific substrates to bind to enzyme
- Bonds between substrate and active site:
Ionic, Hydrogen, Hydrophobic Interactions
Induced Fit:
- Active site no rigid
- Shape of active site changes when substrate binds at active site
- Enzyme keeps substrate under stress
Enzymes ARE MOST ADAPTED to BIND the TRANSITION STATE between SUBSTRATE + PRODUCT
FACILITATES the FORMATION of the TRANSITION STATE
Hence DECREASE ACTIVATION ENERGY of REACTION
Lock and Key:
- Substrate fit precisely into active site
- Substrates that don’t fit won’t be catalysed
Amino acid becomes slightly deprotoated, more based, now charged
Performs NEUROPHILLIC ATTACK on carbon
Pushes e- towards the O, forms transient bond
This creates high energy state
Polar charge on H help neutralise charge, hence stable reaction
Then electrons are pushed again to break the peptide bond
Catalysis Reactions:
- 3 major types:
Acid base
Stabilization of charged intermediates
Providing highly reactive nucleophillic groups
ENZYME CATALYSIS
- Gibbs Free Energy
Gibbs Free Energy:
- Is the potential energy of a chemical system
- Enzyme is stable, then undergoes catalysis, in which it becomes more unstable as free energy increases
- Transition state = Ea
- Subsequently release energy to form products
With enzyme:
- Enzyme create environment with a modified transition state
- This transition state has much lower activation energy, hence reaction needs less time to create products
_ Overall energy is the SAME
ENZYME CATALYSIS
- Catalytic Groups
- Prosthetic Groups
Some enzymes = simple proteins
= only constituents = amino acids
- Groups required for substrate binding + catalysis are provided by amino acid residues
Catalytic Groups of Proteins:
- Use to catalyse reactions using amino acid residues which donate/accept H
Other enzymes = constituents other than amino acid to provide reactive groups at site
- Are bonded covalently/noncovalently to enzyme to contribute extra types of chemistr (eg. coenzyme)
Prosthetic Groups:
- Not all catalytic groups can help with all reactions in enzymes
- Prosthetic groups help out with this
- Non protein group forming part of/combined with protein (collaborate with protein to catalyse reactions)
ENZYME CATALYSIS
- Measurement of Enzyme Activity
- Kinetics of enzyme substrate reactions: Km, V max
- Michaelis-Menten Equation
- Lineweaver-Burk plots
To characterize enzyme function, need to measure the rate of enzyme catalysis in terms of
1. Substrate disappearance
OR
2. Product formation
- Vmax = very efficient enzyme, highest velocity of reaction
- Km = value at which [S] gives 1/2 Vmax
large Km = inefficient
REMAINS CONSTANT NO MATTER WHAT - Low [S], many unoccupied enzymes
- as [S] increases, enzymes occupied, V max slowly becomes constant, until [S] is used up, plateau forms
Michaelis Menten Equation:
E + S ES E + P
Vzero = Vmax/(1 + Km/[S])
Lineweaver-Burk plots:
- Obey Michaelis Menten equation
- Graphical representation of enzyme kinetics
ENZYME CATALYSIS
- Inhibitors and their Lineweaver-Burk plots
Irreversible inhibition:
- Combine with enzyme covalently
- Give a permanently non-reactive form of enzyme
Reversible inhibition
- Combine with enzyme reversibly
- Can regain full activity if inhibitor removed
a) Competitive inhibition - inhibitor binds to active site
- can be reversed by increaseing the concentration of substrate
- Vmax remains unchanged when [S] is increased
Same y, different x and gradient
same 1/Vmax (y-axis)
increase in apparent Km
(x-axis change)
increase gradient as alpha and [I] increase
- efficiency of enzyme interaction with substrate decrease
- as it will be reached eventually with high [S]
b) Uncompetitive inhibition
- inhibitor binds to separate site
- only binds to ES complex, not enzyme alone
- restrict substrate from being processed
- affects both Vmax and Km
Different x and y, same gradient:
different 1/Vmax
Km decreases
- Vmax decreases with increased inhibition as catalysis is prevented
- inhibitor effectively locks the ES complex together
same gradient for all
c) Mixed (non-comp) inhibition
- inhibitor binds to a site other than the active site
- may bind to ES or E complex
- may not affect the binding of substrate
- affects Vmax, not Km
Different y, x and gradient:
different 1/Vmax
gradient increase as [I] increase
- Vmax decreases with increased [I] as catalysis
is prevented
- Effects on Km = combination of interference by inhibitor locking E-S complex
ENZYME CATALYSIS
- Allosteric Enzymes
- Isoenzymes
Enzymes cannot be at 100% efficiency all the time, hence their efficiency is modulated, primarily due to 4 structure and flexibility.
Made up of catalytic site (where active site is) and regulatory domain (were modulator is).
They can exist in 2 different shapes. \+ve Modulation = parallelogram conformation - becomes active -ve Modulation = rectangular - becomes not active
Allosteric Enzymes:
- Characterized by quaternary structure/multi-domain
- Kinetics are different from Michaelis - Menten “model” enzymes (sigmoid kinetics)
- Substrate or modular binding induces conformational (structure) change to facilitate (allow) substrate binding
- Enzyme is flexible
- Cooperative interaction
Side note:
The rate behavior of an enzymatic reaction that yields a sigmoid (S-shaped) curve for a plot of reaction velocity versus substrate concentration
Isoenzymes:
- Have different 1 structure
- Can be separated by a number of physiochemical techniques
- Catalyse same reaction
- Are products of different genes in single cell/tissue type
- Readily measured in biochemistry labs
