Midterm 1 Flashcards
Biochemistry
the study of chemical substances and the vital processes occurring in a living organism
~___ biomolecules within a cell
200
Biomolecule
an organic compound normally present as an essential component of living organisms
Types of Biomolecules
- Nucleic Acids
- lipids
- carbohydrates
- proteins
Nucleic Acids
informations storage (DNA/RNA)
Catalysis (ribozymes, ribosomes)
Energy transfer (ATP)
Components of cofactors such as NAD and FAD
lipids
Barriers (membranes)
Long term energy storage (fatty acids - triacyl glycerols)
Signaling (steroid hormones)
Insulation (blubber)
carbohydrates
Energy and energy storage (glucose, glycogen)
Cell recognition (glycosylation - sugars attached)
Structural (cellulose) (e.g. core of wood is sugar- but not able to break down)
Components of DNA and RNA (deoxyribose/ribose)
proteins
enzymes/catalysis (alcohol dehydrogenase)
Movement (actin & myosin)
Transport (hemoglobin)
Storage (ferritin)
Structural (keratin)
Defense (antibodies)
Regulation signaling/hormonal signaling (insulin - injected to retain protein structure)
Most biomolecules are composed of the following elements
carbon, nitrogen, oxygen, hydrogen
And elements to a Less extent: phosphorus and sulfur
biomolecules interact with each other and themselves by
bonding (different types of interactions) or molecular interactions
5 major types of interactions/bonding
1. Covalent bonds Noncovalent interactions: 2. Hydrophobic interactions 3. Van der Waals interactions 4. Ionic interaction 5. Hydrogen bonds
Covalent bonds
sharing of electrons between 2 adjacent atoms
short solid lines
Tend to be short
hard to break (strong)
Not easily reversible (stable)
High energy - small bond length
Resonance structures
CB bind atoms together to form biomolecules (e.g. adenine)
Can rotate above the single bonds (important for folding)
Geometry of carbon bonding with 4 single bonds
it adapts tetrahedral structure (109.5°) with free rotation about each bond
Geometry of carbon bonding with double bond
When carbon has a Double bond - adopts trigonal planar structure (120°) - no free rotation about double bond and a series of atoms are locked in a plane
Noncovalent interactions are all ____, meaning
electrostatic, meaning there are stationary or partial charges (allow for biomolecules to interact with each other and themselves -e.g. DNA structure, DNA replication, protein folding, and substrate folding)
_____ ______ of the phosphate groups in the DNA backbone
Electrostatic repulsion
Hydrogen bond
a hydrogen atoms partly (unequally) shared by 2 electronegative atoms
(with H donor + and H acceptor -)
This is a special form of an electrostatic interaction
H-bonds are weak (4-13kJ/mole) and longer (1.4 -2.6Å) than covalent bonds
Hydrophobic effect
Dispersion of lipids in H2O - each lipid molecule forces surrounding H2O molecules to become highly ordered
Cluster of lipid molecules: only lipid portions at the edge of the cluster force the ordering of water. Fewer H2O molecules are ordered and entropy is increased
Common H-bonds in biological systems
Bases in DNA can form H-bonds to form base pairs -> H-bonds in DNA base (A:T) pair
Water: Oxygen is partially negatively charged; Hydrogen is partially positively charged
Water’s structure allows for the formation of multiple hydrogen bonds
Water can disrupt H-bonding
Between neutral groups
Between peptide groups
Hydrogen donor
electronegative and tends to pull the electrons away from the hydrogen
The acceptor is also electronegative and thus has a partial negative charge and must have a lone pair of electrons
Cl and F does not act in biological systems but Negatively charged atoms can be ______ ______
electron donors
Ionic interactions
interaction of 2 charged atoms based on coulomb’s law (rolled into dielectric constant)
Attraction
Repulsion
F =q1q2/εr2
what are the variables?
Where F is the force of the interaction
ε is the dielectric constant
Takes into account the medium the atoms are in
In biological systems the atoms are in water and water has a high dielectric constant
Negatively charged backbone of DNA repel each other
Hydrophobic interactions (special type of interaction)
Properties of water and thermodynamics: entropy driven event
Water will decrease entropy (randomness) when a nonpolar molecule is in an aqueous environment
When a nonpolar molecules (hydrophobic) is added to water, the water molecules are forced into a shell (cage) around the molecule (they can’t interact with it-> lowers entropy
When 2 non-polar molecules come together, fewer water molecules are needed to form a shell, and entropy increases (even though the hydrophobic molecules are clustering together)
Favors nonpolar molecules coming together
Nonpolar molecules cause water to surround the molecule in an ordered manner
Hydrophobic molecules coming together -> leads to
decrease in entropy, but the water molecules are becoming more disordered -> increase in entropy
Even though the entropy of the liquids has decreased, the entropy of the water molecules has increased by a greater amount
Van der Waals interactions
attraction of any 2 atoms in close proximity
- a specific form of an electrostatic interaction
Van der Waals interactions in DNA
At any given time, the charge distribution around an atom is not symmetric
That asymmetry causes complimentary asymmetry in other atoms resulting in the 2 atoms being attracted to each other
Van der waals Contact distance- distance between two atoms of maximal attractions between 2 atoms - overlapping electron cloud
The attraction increases until the 2 atoms electron clouds start to repel
Van der Waals interactions in DNA - maximal attraction between
Weak interactions are weak with 2-4 kJ/mole
importance of water
Almost all biochemical reactions/processes occur in an aqueous solvent
Water plays a huge role in these reactions or interactions
Water is cohesive - can interact with itself and anything with charges or partial charges
Water has a bent shape making the molecule polar and capable of forming multiple H-bonds
Water is an excellent solvent for polar (hydrophilic) molecules
Water can form up to hydrogen bonds
4
Amphipathic molecule
hydrophilic (water loving) and hydrophobic (water fearing) properties; molecules that contain polar and nonpolar groups
Water molecules can weaken electrostatic interactions by …
competing for their charge
effects of water in relation to dielectric constant, and H bonds
Water molecules can weaken electrostatic interactions by competing for their charge
Water reduces electrostatic interaction by ~80x
Water has a high dielectric constant
This has serious consequences for biological systems - often water needs to be excluded (or manipulated to allow the various electrostatic interactions to occur
Water is needed to dissolve thing but can also interfere with electrostatic interactions
why must Water be removed for DNA formation
Water must be removed for DNA formation for H-bonds to form between base pairs
Water can disrupt H-bonds by forming its own H-bonds
Thermodynamics laws:
- All biological events are governed by a series of physical laws
1. The total energy of a system (matter in a defined space), and the surroundings is constant
2. The total entropy of a system and its surrounding always increases for a spontaneous process
explain: The total energy of a system (matter in a defined space), and the surroundings is constant
cannot create or destroy energy- can only change its form (e.g. burning wood - converting chemical energy into heat and sound, or dropping ball- PE to KE)
Enthalpy: H:
heat content of the system and its surroundings
- When heat is released, makes the universe more disordered but the reaction is ordered (decrease in entropy)
Entropy: S
measure of randomness
It always increases for a spontaneous process
explain: The total entropy of a system and its surrounding always increases for a spontaneous process
and 2 examples
For any spontaneous process increase E.g. increase in entropy mixing 2 strands of complementary DNA
But things can become ordered. Entropy can be decreased locally in the formation of ordered structures only if the entropy of the universe is increased by an equal or greater amount
E.g. of annealing of 2 strands of complementary DNA; entropy decreases (2 molecules to 1 molecule) but heat is released, causing the entropy of universe to increase
randomness
Using △Hsys and △Ssyst: △G = △H - T△S
what are the variables and what is it used for
T is temperature in Kelvins
Where △G is known as the Gibbs Free energy, measured in kJ/mole
Used to determine spontaneity of a reaction spontaneously: how likely the reaction is to occur
Spontaneity doesn’t define how ___ a reaction will occur, just that it …
fast; can (or cannot occur)
If △G < 0
reaction is spontaneous (exergonic reaction)
- when △H < 0, △G becomes more negative and the reaction becomes more spontaneous
- when △S > 0 (i.e. the reaction becomes more disordered), △G becomes more negative, and the reaction becomes more spontaneous
If △G > 0
reaction is nonspontaneous (endergonic reaction)
__ is very useful because all reactions in a biological system must be spontaneous to occur
△G
The concentration of H ions (i.e. protons) within biological systems is crucial because most biomolecules can act as…
weak acids or bases (i.e. their groups can become ionized)
pH alters the_____ ___ of an amino acid
ionization state;
how a molecules is charged, depends on __ of solution
pH
how does pH affects the ionization state of nucleotides
Need to maintain pH in biological systems
[H+] affects enzyme mechanisms, H-bonding, protein folding
Molecules behavior depends on its ionization state and [H+] affects ionization state
Ionization state
whether a molecules is charged or uncharged
pH
[H+] is measured as pH
pH = -log[H+]
Scale of 0 -14 with 0 being highly acidic and 14 being strongly basic
what is Kw
ionization constant = 1 x 10^14 = [H+][OH-]
Weak acids
acid that is not completely ionized in solution
E.g. (acetic acid) CH3COOH ⇌ CH3COO- (acetate) + H+
COOH = acid; COO- = -ate
Strong acids
acids that is completely ionized in solution
Acid
proton donor
Each acid has its own tendency to lose a proton easily, which is defined by Ka (equilibrium constant)
Henderson-Hasselbalch equation
pH = pKa + log([A-]/[HA])
When the pH = pKa, then [HA] = [A-] (pH =pKa =4.76)
When the pH < pKa, then [HA] > [A-]
When the pH > pKa, then [HA] < [A-]
- We can calculate the pH of any solution if the molar ratio of acid to conjugate base and pKa is known through the Henderson-Hasselbalch equation
Base
proton acceptor
Titration curve of acetic acid
If we titrate a weak acid with a strong base (e.g. NaOH), the acid is releasing protons
If you are adding strong acid, the conjugate base is absorbing the protons
Y-axis = pH
X-axis = OH- added
Titration curve - The curve levels off ->
buffer, where the pH does not change very much even though large amounts of base (or acid) are being added
Equal concentration of acid and conjugate base
In this range the pH does not change much
The buffering range is usually +/-1 (for acetic acid: 3.8 - 5.8)
when do Buffers fail
Buffers fail when you run out of the acid or conjugate base
In the direction of adding NaoH -> when you run about of acid
buffer
Buffer is mixture of acid and conjugate base, resisting changes in pH because both forms are present, and it is most effective when the pH =Pka
Buffers are a storage area for protons
In biological systems, 3 key buffers
- Carbonate buffer system
- Phosphate buffer
- Histidine and cysteine can also buffer
Protein characteristics
a linear polymer built out of amino acids (ɑɑ)
A protein’s final 3-D shape depends on its sequence of aa
Aa contain a large number of different functional groups allowing for massive diversity
Proteins can interact with each other and other molecules to form complexes
Proteins can be flexible or rigid
The Amino Acids
Amino acids contain a Central carbon (alpha carbon) attached to an amino group, a carboxyl group, a single hydrogen group, and a unique side chain “R”
Note that the alpha carbon is chiral
There are 2 enantiomers of each amino acid (except glycine)
Chiral center
an atom with its substituents arranged so that the molecule is NOT superimposable on its mirror image
Cannot overlap mirror image
Enantiomer:
a pair of molecules each with one or more chiral centers that are mirror images of each other
We can draw each aa in the L or D form
In biological systems, almost all aa exist in the _ form (both free and in proteins)
L
L and D form structure
In Fischer projections, if you position the carboxyl group above the C-alpha and the R group below, then the amino group will be left of C-alpha in the L designation and right of C-alpha in the D designation
Zwitterionic aa
has both positive and negative charge
Positive amino; negative carboxyl
the predominant form of aa in biological systems (neutral pH 7)
Assume pH 7 when not noted
There are __ common key aa and these (or slight modification are used in all living things)
20
Nonpolar, aliphatic R groups
Hydrocarbon side chains that are open or nonaromatic ring Alanine (Ala, A) Valine (Val, V) Leucine (Leu, L) Isoleucine (Ile, I) Methionine (Met, M) Proline (Pro, P) Glycine (Gly, G) LIMP VAG
Aliphatic
compound with open-chain structure (alkane)
Glycine (Gly, G)
not really hydrophilic or hydrophobic, not really anything but hydrophobic aa are the best fit
Simplest aa
R = Hydrogen
Only achiral aa
Alanine (Ala, A)
Contains a methyl group CH3
Valine (Val, V)
peace sign/V with CH, and two CH3
Hydrocarbon side chain
Leucine (Leu, L)
Y shape with CH2-CH-(two)CH3
Hydrocarbon side chain
Isoleucine (Ile, I)
has a second chiral center only one form though
Hydrocarbon side chain
Methionine (Met, M)
the start aa when synthesizing protein (unless cut off)
Hydrocarbon sidechain, except it has a nonpolar thioether (CH2-CH2-S-CH3)
Proline (Pro, P)
R group bound to amino group (formed a 5 membered group)
The steric hindrance causes (both cis and trans occur)kinks in the chain
Aliphatic side chain but with a twist, the end of the R group is bonded to the Nitrogen in the amino group
The ring structure makes it more restrained - not that flexible (cannot twist)
often introduces kinks into aa chains i.e. polypeptides
All of the above are hydrophobic and will tend to cluster together -> effect on R groups
Different sized R groups allow for close packing
Often found in the center of proteins (away from water)
Not reactive (more structural roles or binding)
Aromatic amino acids
Contain aromatic rings (e.g. phenyl rings)
Also participates in hydrophobic interactions
Tend to find clustered inside away from water
Phenylalanine (Phe, F)
Tyrosine (Tyr, Y)
Tryptophan (Trp, W)
FYW: Fuck you want
Phenylalanine (Phe, F)
Alanine with a hydrophobic phenyl group
Tyrosine (Tyr, Y)
Like F, but has a reactive -OH group attached to phenol ring that can form H-bonds
has polar groups; more towards the surface near water
The ring makes them predominantly hydrophobic
Typically pKa 10.9
Tryptophan (Trp, W)
5 membered wing and 6 membered ring
5 membered ring has NH on the right that does not lose H easily
Contains an indole group, which are 2 fused rings with an NH group
Reactivity: F < W < Y
Less hydrophobic than Phe
Positively charged Basic R groups aa
Lysine (Lys, K):
Arginine (Arg, R)
Histidine (His, H)
Kiss Right Here
Lysine (Lys, K):
Lysine discovered when they watched cells die and saw lysate come out
Assigned K because, L was taken and they went 1 letter back
4 CH2 then attached to NH3+ at the end
Amino group pKa is high -> greater than 10.8
Stays positively charged at pH 7
Does not like to give up proton -> basic
Contain long chains with ionizable groups (amino group)
Arginine (Arg, R)
pKa above 12
Contain long chains with ionizable groups (quandinium group)
Positively charged at pH 7
Histidine (His, H)
Uncharged at pH 7
pKa is 6 -> close to 7
Can act as a buffer
Has ionizable group (imidazole ring)
It can either be charged or uncharged depending on its location in a proteins
Often found in the active site of enzymes when it can act as a hydrogen donor or acceptor
Easy to manipulate histidine as it is close to 7
Depending on the location change pH -> true for all aa
Charged (groups basic and acidic amino acids) are often found on the surface of proteins (interacting with water and away from hydrophobic aa)
Acidic Negatively charged R groups
Contain carboxylic acids (carboxyl groups) in the R group
Negatively charged at pH 7 (pKa < 4)
Aspartate (Asp, D)
Glutamate (Glu, E)
Polar uncharged R groups
Reactive and capable of h-bonding Not charged More hydrophilic More reactive Serine (Ser, S) Theranine (Thr, T) Cysteine (Cys, C) Asparagine (asn, N) Glutamine (Gln, Q) Queen Can Never Shit Today
Serine (Ser, S)
-CH2OH Contain alipathic (open chain) hydroxyl groups
Theranine (Thr, T)
Contain alipathic hydroxyl groups
Cysteine (Cys, C)
Polar; Can weakly H-bond
Contains sulfidyl (thiol) -SH group
-CH2-SH
Two cysteine comes together -> disulfide bonds
Can link two chains or two parts of the same chain together
Done by oxidation (loss of electrons) of 2 cysteines to a single nonpolar cysteine
Disulfide bond is a covalent bond
E.g. primary structure of insulin
pKa 8.3
Asparagine (asn, N)
Resembles aspartate except instead of -OH it has a H2N
Derivative of aspartate with Nitrogen attached
Cannot ionize the group
Contain terminal carboxyamid instead of carboxyl group
Terminal NH2 is not charged and will not lose its Hydrogen
Glutamine (Gln, Q)
Derivative of glutamate with NH2
Terminal NH2 is not charged and will not lose its Hydrogen
Contain terminal carboxy amid instead of carboxyl group
Terminal alpha carbonyl group typical pKa
3.1
Aspartate (asp, D)
-CH2 - COO-
Acidic: low pKa
aspartic acid pKa 4.1
Glutamate (Glu, E)
-CH2 - CH2- COO-
Acidic: low pKa
Glutamic acid pKa 4.1
Terminal alpha amino group typical pKa
8.0
Protein structure
Primary structure 1° -> secondary structure 2° -> tertiary structure 3° -> quaternary structure 4°
Primary structure
linear sequence of aa linked by peptide bonds to form a protein
Protein synthesis in ribosome
condensation reaction
Spontaneity wise -> formation of protein is not favourable
Must add energy to for it to form
High activation energy required to break peptide bond
Peptide bond is polar -> polypeptide is polar
Peptide bond
linkage of an alpha carbonyl of on aa to the amino group of another aa with the loss of water
Not energetically favourable but once formed is stable
Polypeptide
series of aa residues linked by peptide bonds Polar -> have 2 different ends Free amino end (-NH3+) -> N terminal end Left side Involved in ionic interaction Free carboxyl end (-COO-) -> C terminal end Right side Involved in ionic interaction
backbone
(repeating unit N-C(a)-C) and unique R group
Backbone is hydrophilic and can form H-bond with each group capable of doing one H-bond
All the carbonyls and N-H groups in the backbone can hydrogen bond (The C=O and N-H can each form 1 hydrogen bond
Important for forming 3D structures)
Exception: proline
why does proline have limited H-bonding
the R group is connected to the N of the amino group so there is no H attached to the N
Residue
aa unit in a polypeptide
Daltons (Da)
Weight of proteins is expressed in Daltons (Da) or more commonly kDa
Same as molecular weight
1 Da = mass of H atom ~ 1g/mole
Note about nomenclature
A point mutation is usually written as original aa location and new aa
D 614 C =
D 614 C = aspartate at position 614 has been replaced by cysteine
Knowing the 1° aa sequence should help us to
- Determine shape: 3D shape of protein depends on its sequence of aa
- Understand function
- Understand disease (e.g. within aa are binding to covid)
- Understand evolutionary history (compare sequences of protein and see how they changed over time)
explain how proteins are flexible but conformationally restrained
Backbone of a polypeptide is restrained due to double bond characteristics of the peptide bond
Because of resonance between the peptide bond and the carbonyl group
Result: peptide bond is planar and locks a series of atoms into a plane
There is no rotation about the peptide bond
The carbonyl oxygen has a partial negative charge and the amide nitrogen a partial positive charged, setting up a small electric dipole
configuration about the peptide bond
Peptide bonds could technically exist in cis or trans as the peptide bond acts as a double bond
Virtually all peptide bonds in proteins occur in the trans configuration
except X (aa)-pro (proline) (both cis and trans occur)
Cis configuration has steric hinderance
More correct in zigzag fashion -> trans configuration
dihedral angle (aka torsion angle)
We can measure the amount of rotation about the bond
Ranges from -180° to +180°
The bond between N-C and the bond between C-C=O are free to rotate
This provides flexibility allowing the protein to fold in many different ways
Dihedral angles
The N-C(a) dihedral angle = ɸ(phi)
The C(a)-C=O dihedral angle = Ѱ (psi)
Not all combinations of angles of Ѱ and ɸ are permissible
due to steric hindrance
Further limits the number of structures a protein can adopt
Ramachandran plot
combinations of dihedral angles that are permissible are shown in this plot
This plots is the same for 18 of the aa (different from proline and glycine)
Proline considered alpha helix wrecker
Areas of dark blue are favourable
Areas of light blue are borderline
Note that ¾ of all angle combinations are not possible (white)
Large molecules that can freely rotate among many bonds will adopt ____ ___(i.e. a mixture of many different structures)
random coils
They can often spontaneously fold into a single structure under ______ ____
physiological conditions
Secondary structure (2°)
the spatial arrangement of aa residues that are close to each other in a linear sequence
Alpha ɑ helix:
Beta 𝛃 sheet
Alpha ɑ helix:
polypeptide backbone forms the inner part of a right-handed helix, with the side chains (i.e. R) sticking outwards
The helix is stabilized by intrastrand hydrogen bonds between the NH and C=O groups of the backbone (N-C-C repeated)
Intrastrand H-bond = H-bonding with itself which helps hold the helix together
is there space in the centre of the helix?
No space in the centre of the helix -> everything is packed together
The helix is stabilized by intrastrand hydrogen bonds between which groups and how many residues away
The carbonyl of residue (i) forms a H-bond with the N-H 4 residues further down the chain- closer to C-terminal (i.e. residue i + 4)
Alpha helices have ideal dihedral angles of
Ѱ= -45 and ɸ = -60
R groups i, i+1, i+2 are pointing
R groups i, i+1, i+2 are pointing away from each other
The different R groups in this proximity a hydrophobic and hydrophilic groups that point away from each other -> amphipathic
R groups i, i+3, i+4 are pointing
R groups i, i+3, i+4 are pointing in a similar direction
Each amino acid residue in the helix increases the helix length by
1.5 Å (i.e. we say the helix rises by 1.5Å)
Left-handed helices
left hand helices are permitted but rare as they are not as stable due to the fact that the amino acids are in L
Alpha helices are depicted as
twisted ribbons or a rod
Usually the maximum length of a helix are ___ or less
45 Å or less
keratin structure
keratin are 2 alpha-helices can intertwine into coiled coils
Beta barrels are depicted as _______ _____
twisted arrows
in a parallel B sheet, each aa residue extends a B-strand by ____ Å
3.25
Tertiary Structure (3°)
The spatial arrangement of aa residues that are far apart from each other in linear sequence as well as the pattern of disulfide bonds
IMPORTANT because 3° is the 3D structure
myoglobin
O2 storage protein in mammalian muscles Red in steak is not blood (hemoglobin) -> myoglobin (O2) capacity Globular protein with no symmetry Very few voids (i.e. holes in the core) Single polypeptide chain of 153 aa
what does the heme group in myglobin do?
heme group (ison in a protoporphyrin ring)
Where the O2 binds
In myoglobin the O2 binds tightly and is only released when [O2] is low
how many a-helices in myoglobin
70% of the chain is in a-helices (8 a-helices)
Most of the rest is in loops and turns
The core of the myoglobin protein is almost exclusively composed of
The core of the protein is almost exclusively composed of hydrophobic residues (Except for 2 His which are needed at the O2 binding site)
structural domains
Some proteins can have multiple regions called structural domains lined by flexible sections of the polypeptide (often with no defined structure)
In most tertiary structure, the dihedral angle for each residue - Ramachandran plot
In most tertiary structure, the dihedral angle for each residue in the protein falls into the permissive area in a Ramachanran plot
Quaternary structure (4°)
Folded on its own
The spatial arrangement of multiple subunits (polypeptides) and the nature of their interactions
Some proteins are composed of more than one polypeptide chain
Some proteins must be multimers (protein with quaternary structure) in order to function
homomers
Quaternary structure (4°) Chains can be identical
Heteromers
Quaternary structure (4°) Chains can be different
Hemoglobin (H6)
O2 transporter in mammals
Hemoglobin can only do its just as a tetramer (composed of 4 subunits: 2 a-subunits and 2-B-subunits)
It is the interactions between the subunits that are critical for function, hemoglobin can’t function unless it is a tetramer
quaternary structure example: the capsid of viruses (in this case, minute virus of mice)
Composed of 9 VP1 and 51 VP2 protein subunits to form an isohedral (polygon with many sides) capsid (complex quaternary structure) with just enough room the fit the viral DNA
how many structures can a protein adopt
Even with limitations on the backbone of a polypeptide, there are trillions of structures it could adopt and it would take forever for a protein to try each one
But, most proteins fold into just one structure in less than a second
Alphafold2 and riseltafold
Up until 2 years ago, we could not predict a protein’s 3D structure based on primary sequence but nature could
Now, computer AI programs such as Alphafold2 and riseltafold
what is folding driven by
Folding is driven by thermodynamics (i.e. finding the most stable complex - most negative △G)
The free energy change between folded and unfolded proteins is small (20-60kJ difference)
This is partly driven by entropy, ie the hydrophobic residues are excluded from water in the core while the hydrophilic residues are on the surface
Many B-sheets and a-helices are ______
amphipathic
In order to bury the backbone of a polypeptide in the core, it needs to _____
H-bond
what can destabilize structures
Unpaired charged or polar groups in the core can destabilize structures
Can a portion of the primary sequence be used to define secondary structure?
Yes and no. certain aa residues are more likely or less likely to be found in and stabilized or unstabilized a-helices and B-sheets
The more positive the delta G value, the more destabilized
aa acids such as Pro and Gly destabilize a-helices
Experiments have shown that the exact same protein sequence in 2 different proteins can adopt 2 different secondary structures
We can’t always determine secondary structure by looking at a portion of a primary structure
Tertiary structure influences 2ndary structure
Protein folding process
Folding is an all or none process, either protein is folded or it is not
what does it mean that proteins can breathe
proteins can breathe: i.e. they can flex and open and close
what does it mean that “Folding is thought to be cooperative”
There may not be just one pathway that a protein follows to fold
I.e. as one one protein of the protein folds (for example, an a-helix) , it will influence how another protein folds - a protein doesn’t have sample every possible structure
There might be multiple pathways to the folded state
Not all proteins have a single proteins structure
Some might only form a structure when bound to another protein
Some proteins exist in an equilibrium between 2 structures
Modifications of aa in proteins (4)
aa can be post translationally (after synthesis of protein) covalently modified
- phosphorylation
- glycosylation
- hydroxylation
- carboxylation
Proteins can also be cleaved and trimmed after synthesis
(in fact most are)
E.g. fibrinogen (inactive) -cut-> fibron (active)
Many viruses make super long polypeptides that are then cut into small functional proteins
Enzyme
biological macromolecule that acts as a catalyst for biochemical reactions - usually proteins
Enzymes are very specific, they will only catalyse are specific set of reactions
Catalyst
a chemical that increases the rate of the reaction without being consumed
Enzymes speed up the rate of reaction - they are essential for biological synthesis
Rate can be read as molecules of substrate coverted per second (per molecule of enzyme if it is present)
proteolytic enzymes (i.e. proteases)
cleave peptide bonds
Trypsin
cleaves only the peptide bond on the carboxyl side of Lys and Arg
Thrombin
cleaves only Arg-gly protein bond
Specificity is based on
a series of weak interactions between the substrate and the enzyme, especially in the active site
The shape of the enzyme (Especially at the active site) determines (2)
specificity and function
Active site
the region of the enzyme that bonds the substrate
It contains the residues that directly participate in the reaction
Characteristics of active sites
Tends to be a cleft or open hollow part for substrate to come in
The residues in an active site can be far apart in the primary sequence and fold to come close together
Cleft in the enzyme made up of residues from all over the primary aa sequence
They take up a small volume of the enzyme
Water is usually excluded from the active site, changing the behaviour of the residues -> hydrophobic effect
The substrate is bound to the active site by a series of weak interactions
is the substrate and enzyme complementary
The substrate and the enzyme must be complementary, otherwise the substrate cannot bind and catalysis cannot occur
The active site is not a perfect complementary fit to the substrate
lock and key
(rare) complementary match to the substrate
too specific and hard for the product to let go of substrate
Induced fit
binding of the substrate causes the active site to assume matching shape
enzyme does have a perfect fit but the enzyme changes shape
Cofactor
an inorganic ion or small organic compound (often referred to as a coenzyme) required for enzymatic activity
The heme requires Fe2+ as cofactor
Prosthetic group
A cofactor that is tightly bound to an enzyme
E.g. myoglobin (tertiary structure) has a heme prosthetic group
FAD and NAD as cofactors
FAD (more like a cofactor than NAD, though both are cofactors) fits more perfectly to enzyme
Apoenzyme
An enzyme without its cofactor
Holoenzyme
an enzyme with its cofactor
Enzyme thermodynamics
- Enzymes do not alter the final equilibrium of products to reactants
- Enzymes do not alter △Grxn -> they obey the laws of thermodynamics
I.e. they can’t change the spontaneity of reaction
If △G of a reaction is (+), its is a nonspontaneous and adding enzyme will NOT change that - Enzymes speed up the rate of reaction
enzymes accelerate reaction by
decreasing the activation energy (△G‡) by facilitating the formation of transition state (X‡)
Imagine substrate being converted to a product
In order to form the products, the substrate goes through a transition state (X‡)
The transition state has the highest G (Gibb’s free energy) in the reaction and the lowest concentration
how does the activation energy controls the reaction rate?
only a fraction of Substrate will have enough energy to form X‡
Activation energy (△G‡) : the energy needed to get to transition state (X‡)
△G‡ (s->p) = G(X‡) - G(s)
△G‡ (p->s) = G(X‡) - G(p)
the activation energy is not part of the over △Grxn calculation because the energy put in is returned when the X‡ is converted to products
Consider enzyme converting substrate to product
S +E ⇌ ES ⇌ EP ⇌ E +P
Enzymes interact with the transition state such that the activation energy is lowered
The reaction speeds up as a greater fraction of Substrate has the energy reach X‡
Where does the energy come from to lower the activation energy?
binding energy (△GB)
Enzymes coming and enzyme binding and stabilizing the transition state
The active site of an enzyme is complementary to the transition state
Binding energy (△GB): the energy derived from the interaction between the enzyme and substrate
Beta 𝛃 sheet
Beta barrels are depicted as twisted arrows
More common because more stable
2 or more 𝛃 strands (polypeptide strands usually from the same molecule) associated as stack of chains in an extended zigzag form stabilised by interstrand hydrogen bonds
The planarity of the peptide bond means different sections of the sheets are fixed in different planes and the dihedral angle set the angle at which those planes intersect -> forming plates
Do not normally see beta-strand by itself
antiparallel beta strands
Each aa residue extended the B-strand by 3.8 Å (more spread out than an a-helix)
The R groups of adjacent residues (in a B-strand) point in opposite direction
The strands are arranged into pleated sheet
In an antiparallel sheet, the NH and C=O of one residue on one strand (i) H-bonds to a single residue (j) on the other B-strand
Antiparallel sheets have ɸ= -139 and Ѱ=+135 (idealised angle)
Twists and loops - Beta sheets
Peptide chains often reverse direction
Can be accomplished by B-turns (have a defined secondary structure or by larger loops (i.e. no common/defined secondary structure)
Parallel B-sheet
H-bonds are weaker
The NH of one residue (i) in one B-strand H-bonds to a C=O on residue (j) on other B-strand
The C=O of residue i H-bond with the NH two residues further (j+2) on the other B-strand
Parallel B-sheets have ideal angles of ɸ= -119 and Ѱ=+113
in a parallel B sheet, each aa residue extends a B-strand by 3.25 Å
Mixed beta sheet
Sheets can be twisted
Beta sheets are depicted as broad areas pointing to the carboxyl terminal (i.e. C-terminal)
The distance in primary aa sequence (structure) between beta strands can be small or large
Small: Can be quick hairpin turn and another B-strand
Large: Or there could be 100s of aa forming other structures between B-sheets
B-strands (and thus B-sheets) can be flat or twisted
phosphorylation
attachment of a phosphate group to the OH of an amino acid (e.g. Ser, Thr, Tyr)
Signal transduction
E,g, phosphoserine
Glycosylation
attached of one or more sugars to a residue (usually Asn, thr, or ser)
Surface labelling
Hydroxylation
the addition of an OH group (usually a proline)
Fibre stabilisation
Hydroxyproline -> becomes polar with addition of OH group
Carboxylation
addition of a carboxyl group to glutamate
E.g. clotting
Carbohydrate-asparagine adduct
Y-carboxyglutamate
