Week 1 Flashcards
True or false:
Organic compounds that form hydrogen bonds with water are not soluble in water
False- if they can form hydrogen bonds they are usually soluble in water
What is the equation for the Ion Product of water?
Kw = [H+][OH-] = 1 x 10 to the -14
at pH of 7 the solution is ______ and both the [H+] and the [OH-] are ______.
Neutral
1x10 to the -7 (equal parts H+ and OH+)
If the concentration of H+ is 1x10-6 , what is the concentration of OH- ?
Kw/[H+} = [OH-]
1x10-14/ 1x10-6 = [OH-]
[OH-] = 1x10-8
What is pH defined as ?
The negative log of the concentration of hydrogen ions in a solution. This determines the acidity of the solution.
what is the equation for pH ?
pH = -log[H+]
Define an acid?
An acid is a proton donor, and chemical that can “lose or donate” a hydrogen ion (proton) in a solution
Define a base?
A proton acceptor, any chemical that can “gain or accept” a Hydrogen ion (proton)
Define the Conjugate base?
The ion or molecule remaining after the acid has lost a proton
Is also referred to as the salt of the acid (often “ate” ending as opposed to “ic”)
Define Strong acids?
- completely dissociate at any pH
2. when added to water, complete dissociation into the conjugate bas and protons
What 3 Strong acids does the body create?
Sulfuric acid
Hydrochloric acid
Nitric Acid
What is the chemical composure of Sulfuric acid?
H2SO4
What is the chemical composure of Hydrochloric acid?
HCl
What is the chemical composure of Nitric Acid?
HNO3
True or false:
Strong Acids can sometimes be used as buffers
False - strong acids cannot for buffers (they always dissociate )
Define Weak acids?
- Partial dissociation into conjugate base and protons
- Ratio of conjugate base (salt) to weak acid can be adjusted by adding H+ or OH- to the solution
- Only weak acids can buffer aqueous solutions
What is the dissociation equation of a weak acid?
HA = H+ + A-
HA is weak acid concentration
A- is conjugate base concentration
H+ is proton concentration
What is the equation for the dissociation constant Ka?
Ka = [H+][A-] / [HA}
What is the relationship between pKa and Ka in equation form?
pKa = -log(Ka)
When the pH is equal to the pKa , what does that mean for the dissociation of the weak acid HA ?
50% dissociation
What is the Henderson-Hasselbalch equation?
pH = pKa + log {[A-]/[HA]}
In relation to the Henderson Hasselbalch equation, when the [A-] is equal to the [HA], what does that infer?
pH = pKa
When you increase the [HA] (acid) in the solution, what does it do?
brings pH down (inverse relationship)
When you increase the [A-] (conjugate base or salt), what does it do to the solution?
brings the pH up (Direct relationship)
What two factors determine the effectiveness of a Buffer?
- the pKa of the buffer relative to pH of solution
2. Concentration of buffer present
When are Buffers most effective?
- When pH of solution is equal to the buffers pKa
2. Generally, when pH of solution is within one pH unit of buffer’s pKa (higher or lower)
What happens when an acid (H+) is added to the buffered solution?
Conjugate base (salt of the buffer) combines with the added acid and prevents pH from changing and turns into HA
What happens when a base (OH-) is added to the buffered solution?
The Acid of the buffer (HA dissociates to donate H+ and A- increases) combines with the OH- and makes water and prevents major changes in pH
Within what range can a buffer compensate for influx or removal of Hydrogen ions?
approx. 1 pH unit of its pKa
After the pH increases or drops more than 1 unit, the ratio of A- to HA changes from 1:1 to 1:10 and the curve becomes exponential
What does the Primary structure of a protein start with?
Written from the Amino terminus (N-terminus) to the carboxyl terminus (C-terminus)
Define the Primary structure of a protein?
A linear sequence of amino acids in polypeptide chain
What is a Zwitterion?
A molecule or ion having separate positively and negatively charged groups (amino acids)
What is the basic structure of all amino acids?
Alpha amino acids:
amino group is attached to alpha carbon (C next to carboxylate group)
Alpha carbon has 2 additional subunits: Hydrogen atom and a R-group
At physiological pH of 7.4, what is the charge in an amino acid for the amino group and the carboxylic acid group?
Amino group = positive
Carboxylic acid = negative
What is the pKa for primary carboxylic acids of all amino acids?
approx. 2 (1.8 to 2.4)
When is the Carboxylic acid protonated ?
When the pH value is much lower than the pKa (high hydrogen ion concentrations are around)
What is the pKa for the amino group?
approx. 9.5 (8.8 to 11.0)
so that at the lower pH of 7.4, most amino groups are fully protonated at carry a positive charge
True or False:
All amino acids are water soluble at pH 7.4?
True
All amino acids are D or L configuration in mammals?
which is the exception?
All are L formation with Amino group on left and Carboxyl group at top
Exception: Glycine - it has 2 Hydrogen groups that makes it neither D or L configuration
Where do amino acids join together?
From one amino group to the carboxylic acid of the next amino acid, thus forming a backbone with the R groups protruding outwards opposite sides every other amino acid
What is the bond between amino acids called? what is released when the bond is formed?
Water is released (Carboxyl gives up one Oxygen and Amino gives up 2 Hydrogens)- Carbon bonds to the Nitrogen
Peptide bond
What is the simplest amino acid?
Glycine - R group is one H atom
Where is Glycine normally found in the chains?
In bends or tightly packed parts of the protein due to is small size
What is unique about the function of Proline?
Contains a ring that involves a connection between the alpha carbon and the alpha amino group which become part of peptide backbone
-this makes a kink in backbone preventing normal configuration
What are the features of non-polar, Aliphatic Amino Acids?
Besides Glycine and Proline
- all others have bulky non-polar R groups
- high degree of hydrophobicity with high hydropathic index because electrons are shared equally between carbon and hydrogen in R chain so they prevent bonding with water
Where do you normally find the non-polar, Aliphatic amino acids?
Hydrophobic rings so they often cluster to form hydrophobic cores
-Van der Waals force present holding them together when many atoms packed closely together
Name the Non-Polar, Aliphatic amino acids (6)?
Glycine Alanine Proline Valine Leucine Isoleucine
gly
Glycine
ala
Alanine
pro
Proline
val
Valine
leu
Leucine
ile
Isoleucine
What are the 3 Aromatic Amino Acids?
Phenylalanine
Tyrosine
Tryptophan
phe
Phenylalanine
tyr
Tyrosine
trp
Tryptophan
What are the 4 Polar, uncharged amino acids?
Asparagine
Glutamine
Serine
Threonine
asn
Asparagine
gln
Glutamine
ser
Serine
thr
Threonine
What are the 2 Sulfur Containing amino acids?
Methionine
Cysteine
met
Methionine
cys
Cysteine
What are the 5 Charged amino acids?
Aspartate Glutamate Arginine Lysine Histidine
asp
Aspartate
glu
Glutamate
arg
Arginine
lys
Lysine
his
Histidine
What do the Aromatic Amino acids have in common?
All have ring structures with similar properties but different polarities
6-membered hydrocarbon ring with 3 conjugated double bonds (benzene ring or phenyl group)
What determines for Aromatic Amino acids if the acid engages in polar or hydrophobic interactions?
The substituents on the ring itself
Describe the ring for Phenylalanine?
Carbon and hydrogen share electrons equally , very nonpolar hydrophobic - rings can stack on each other
Describe the ring for Aromatic tyrosine?
Same ring as Phenylalanine but there is a Hydroxyl group on the ring that can engage in Hydrogen bonds
-more polar and more hydrophilic
Describe the ring for Aromatic Tryptophan?
Complex ring
- indole ring with nitrogen that can engage in hydrogen bonds
- more polar than phenylalanine
Describe the features of the Polar, uncharged group (also classified as Aliphatic) ?
Contain an amide group (asparagine and glutamine)
or Hydroxyl group (serine and threonine)
-such groups allow hydrogen bonds with water, very hydrophilic
-found often on surface of water soluble globular proteins
What does Cysteine play an important role in doing (sulfur containing ) ?
pKa of 8.4 so is usually dissociated at pH of 7.4
- Plays important role in holding 2 polypeptide chains together - disulfide bonds with (SH) Sulfhydryl group
- not very water soluble in blood and tissue
Describe the features of Methionine?
Nonpolar amino acid with large bulky side chain that is hydrophobic
- No SH (sulfhydryl group) to make disulfide bonds
- Important for metabolism to transfer methyl group (CH3) to sulfur atom of other compounds
What charge does Aspartate and Glutamate have at Physiological pH of 7.4 ?
They have Carboxylic acid groups that give them a negative charge at pH 7.4
-Called the Negative or Acidic Charged Amino Acids
What 3 amino acids have side chains containing Nitrogen that can be protonated and positively charged at pH 7.4 and lower pH values? (Basic)
Arginine
Lysine
Histidine
What is different about Histidine than the other Basic (positively charged) polar amino acids ?
Arginine and Lysine do not have imidazole ring for a side chain and have linear chains
What is the pKa for Aspartate ?
3.9 (ionized at pH 7.4 - negative charge)
If pH is less than pKa ?
Protonated (Amino group H3N+ on amino acids)
if pH is more than the pKa ?
Ionized (Carboxylic group COO- )
Which two amino acids are most likely to be dissociated at physiological pH of 7.4?
Aspartate (pKa of 3.9) - Negative
Glutamate (pKa of 4.1) - Negative
-both have Carboxyl groups that lose H+ at pH 7.4
What is the pkA of Histidine?
- 04 (acts as strong base)
- unique because it can be both donor and acceptor of H+ at pH 7.4
What is the pKa of Cysteine?
- 4
- Protonated at pH7.4
What is the pKa of Tyrosine ?
- 5
- Protonated at pH 7.4
What is the pKa of Lysine ?
- 5
- Protonated at pH 7.4
What is the pKa for Glutamate ?
- 1
- Ionized at pH 7.4 (dissociated into A- form)
What is the pKa for Arginine ?
- 5
- Protonated at pH 7.4
For the Polar, uncharged groupings of amino acids
Asparagine, Glutamine, Serine, and Threonine , what is the charge at biological pH of 7.4?
Side chain is protonated
What are the features important to Histidine ?
Strong base for pH 7.4
Can be ligands for Zn or Iron (cofactors)
The two N in the ring can function as nucleophile or electrophile
INTEGRAL for Oxygen binding to hemoglobin
Which 3 amino acids can carry ammonia (toxic) within the body?
Glutamine
Glutamate
Alanine
What is a function of Glutamine (Polar,uncharged) ?
Carries ammonia within the cell
What is a function of Glutamate ?
Neurotransmitter
Used for synthesis of other amino acids
Used to make Glutathine (reducing agent in RBC)
What is the function of Alanine?
Involved in Alanine-glucose shuttle in skeletal muscle
Carries ammonia
Deaminated to form Pyruvate (rapidly shuttles carbon)
What is the function of Serine/Threonine (polar, uncharged) ?
Can be phosphorylated to change the functionality of a protein
What is the function of Tyrosine ?
Can be phosphorylated to change the functionality of a protein
Precursor to many neurotransmitters (Dopamine)
What is the function of Cysteine ?
Forms Disulfide bonds- increases structural integrity
What is the function of Methionine?
Translation start amino acid
Can be metabolized into homocysteine to cysteine
At a pH of 8 , what would the net charge of aspartate be?
-1
When does Post translational modification usually occur ?
Once protein has already folded into its 3D conformation
Protein kinases phosphorylate proteins only at certain hydroxyl groups on amino acids side chains. Which group of amino acids are the only ones that contain hydroxyl side groups?
Serine/Threonine
and
Tyrosine (has a aromatic ring where OH- attaches)
G
Glycine
A
Alanine
P
Proline
V
Valine
L
Leucine
I
Isoleucine
F
Phenylalanine
Y
Tyrosine
W
Tryptophan
N
Asparagine
Q
Glutamine
S
Serine
T
Threonine
M
Methionine
C
Cysteine
D
Aspartate
E
Glutamate
R
Arginine
K
Lysine
H
Histidine
What 3 amino acids can be phosphorylated because they have OH- (hydroxyl groups) in their side chains?
Serine/Threonine
and
Tyrosine (precursor to neurotransmitters)
Which amino acid can form disulfide bonds to increase integrity of peptide chains?
Cysteine
What is the Kd of water ?
What is the Kw?
Kd is 1.8 x 10-16
Kw is 10-14
What are possible post-translational modifications on amino acids?
Glycosylation Fatty Acylation or Prenylation Regulatory Modifications (phosphorylation, acetylation, ADP-ribosylation ) Carboxylation Oxidation
What is Glycosylation ?
Oligosaccharides (small Carbohydrate chains) are bound to proteins by N links or O links
N links- protect from proteolysis or immune attack
O-links = attack glycogen (or others) to proteins intracellularly or secreted on the Hydroxyl group or Serine or Threonine
What does Fatty Acylation or Prenylation do to proteins?
Adds a Hydrophobic (fatty acid) compound to a protein often on lipid membranes
-typically Cysteine residue and involved in regulation
What does phosophorylation to to proteins and who can it bind to ?
Kinase used to bind it to -OH group on:
Serine, Threonine, or Tyrosine
-introduces a large bulky negative charged group that alters function
What is Acetylation do ? what does Acetyl look like?
CH3CO
- transfer from one molecule to another - like Acetyl-CoA
- On the NH2 terminus of Lysine
What does ADP-ribosylation do ?
Transfer of ADP-ribose from NAD+ to an arginine, glutamine, or cysteine residue on target membrane
-primarily in leukocytes, skeletal muscle, brain,testes
How is the alpha helix bond formation formed to make the helix?
Each Oxygen of a carbonyl group forms a hydrogen bond with the hydrogen atom attached to a Nitrogen atom 4 amino acids farther down the chain
-highly compact and rigid
How is tertiary structure defined ?
Association of secondary motifs or structures
- Actin Fold (binds ATP)
- Nucleotide binding fold (only one subdomain will bind NAD)
What is an Amyloid?
Plaque where the native fold was destabilized and then they protein aggregates in Fibrous way burying hydrophobic portions of protein - creates large Fibers (plaques)
How many alpha helix are there in the tertiary structure of myoglobin?
8
What is a prosthetic group? example?
Permanently bound to the protein
- required for function
- Example is Heme bound to Myoglobin and Hemoglobin- used to bind Oxygen
What is a Holoprotein ?
The prosthetic group is bound with its Apoprotein
What is a Apoprotein?
The protein that together with a prosthetic group makes the Holoprotein for a certain function
Describe the binding curve of Myoglobin to O2?
Hyperbolic shape
-Saturation with oxygen very quickly with little pressure of O2 required
What is Myoglobin good for?
Usage in anaerobic activity because of its tight binding to Oxygen where it will only release under extreme conditions - usage in muscles
How many Heme groups does each Myoglobin have?
What type of binding?
1 Heme group
-Non-cooperative
What shape is the binding curve for Hemoglobin?
Sigmoidal shape
- Positive cooperativity
- Must have quaternary structure to display cooperative binding
Describe the Ka vs the Kd for Myoglobin?
High Ka- holds tightly to Oxygen
Low Kd- very hard to dissociate
What partial pressure of oxygen (mm HG) does Myoglobin become 50% saturated with Oxygen?
P = 2.8mm HG
What partial pressure of oxygen (mm HG) does Hemoglobin become 50% saturated with Oxygen?
P = 26 mm HG
vs Myoglobin which is only 2.8mmHG
When the concentration (pressure) of oxygen is high, like in the lungs _______.
Both Myoglobin and Hemoglobin are saturated
Hemoglobin is formed from 4 subunits and is called a ____.
What types of subunits?
Tetramer
-2 alpha and 2 beta subunits (each with its own Heme)
In what state is the affinity for Oxygen low for Hemoglobin?
T state (tense state)
In what state is the affinity for Oxygen high for Hemoglobin?
R state (relaxed state)
What amino acid holds the Iron (Fe2+) group in place just outside of the porphyrin ring of Heme?
Proximal Histidine
Fe is bound in total to 4 Nitrogen atoms in center of heme porphyrin ring
What happens when O2 binds to the Fe2+ in Heme?
The Iron is pulled into the porphyrin ring and energy is used to break the salt bridge previously holding the Iron back
-leads to change in helix shape of ring - one movement making each further movement easier to break (to bind more Oxygen)
Where does CO2 bind to Hemoglobin (Hb)?
at N-terminus and lysine amino
What happens when CO2 binds to Hb?
Decreases affinity of Hb for O2
What can cause the release of O2 from Hb?
Drop in pH (increase in CO2 will cause this in working muscles or ATP production)
- working muscles produce H+ (increase acidity)
- working muscles require more O2
WHat is the Bohr effect?
Physiological phenomenon that Hemoglolins affinity for O2 binding is inversely proportional to the level of acidity and carbon dioxide concentration
What happens to the curve of Oxygen affinity for Hemoglobin as pH drops ?
What is the technical name for this phenomenon?
As pH drops, the curve becomes less sigmoidal and less steep in slope - more pressure or O2 is needed to keep the same level of saturation - hence Oxygen has less affinity as pH drops and is released
(pH is an Allosteric effector of O2 binding)
pH is an ______ _____ of O2 binding on Hemoglobin.
Allosteric Effector (example of linked binding)
What is an example of secondary modifications of proteins to hemoglobin?
Glycosylation of Hemoglobin
- Addition of Glucose to globin chains (protein that surrounds heme)
- can be quantified - HgA1C - used to manage diabetes
What do all amino acids contain?
Alpha Carbon Carboxylic acid Amino group Hydrogen Atom R- group (determines function)
What 3 things is the body normally buffered by?
Proteins
Bicarbonate
Phosphate
Describe a globular structure protein?
Soluble and are rounded
Hydrophobic amino acid core, Dense, Van der Waal applies
Charged polar amino acid on surface- forms salt bridges and ionic interactions
DNA binding
Describe Transmembrane structure protein?
Span cell membrane
Have both hydrophobic and hydrophilic properties
Rigid alpha helixes connected by loops
Usaully have post translational modifications
Describe Fibrous structure protein?
Repetitious subunits
Usually for matrix integrity
Collagen is a good example
Where is Hemoglobin found and Myoglobin?
Hb = travels in blood and transports Oxygen Myoglobin = stays in heart and skeletal muscle to bind oxygen released by Hb
What 3 things stabalize Hemoglobin and Myoglobin that are Homologous proteins?
Tertiary and quanternary structures: Hydrophobic interaction Hydrogen Bonds Salt bonds (weaker in presence of water)
What is the structure of Myoglobin?
8 alpha helixes linked together by alpha turns
Monomer
Hydrophobic pocket containing heme with a ferrous iron atom (Fe+2) at center for oxygen binding
Why is Fe2+ always bound to a Histidine R group ?
Stabilizes the reduced state of iron when it binds to oxygen
How is HbA1c made in the blood?
Spontaneously from Hb and glucose
bound non-enzymatically and irreversibly to amino acid terminus of beta chain of Hb- reliable to give concentration of glucose in blood
What does HbA1c measure ? time frame/
what level is considered pathological?
Measures average glucose in the blood for past 6-8 weeks
level above 5.2% is considered pathological
How does an enzyme function in lowering the activation energy of reactions?
Lowering the activation barrier between reactants and the transition state, thereby increasing the fraction of reactants able to achieve the transition state
- Stabalize the normal transition state
- Alternative pathway from reactants to products
What 4 strategies do enzymes use to reduce the transition energy(top of curve at height of activation energy?
- Transition state stabilization by induced fit
- Acid-base catalysis
- Covalent catalysis
- Meta ion catalysis
What is a carbonyl group?
C=O
When is general acid-base catalysis used?
Method to lower activation energy.
Hydrolysis of ester/peptide bonds, phosphate group reactions, addition to carbonyl groups, etc.
True or false:
Enzymes do not avoid unstable charged intermediates in reaction (which would have high free energy).
False - they do have groups appropriate for :
- donating a proton (act as general acid)
- accept a proton (act as general base)
In chymotrypsin, the active site histidine acts as a ____ to deprontonate an adjacent serine residue
Base
What is a nucleophile?
Donates a proton (H+) to an electrophile
What is covalent catalysis ?
Method used to lower activation energy.
Transient formation of a catalyst-substrate covalent bond
What are the 4 biologically important nucleophilic groups for covalent catalysis ?
Hydroxyl group (OH) Sufhydryl group (SH) Amino group (NH3) Imidazole group (3 carbon ring with 2 NH points in ring- 1 H dissociates)
Which side chains can act to donate protons?
Aspartic Acid
Glutamic Acid
(Histidine sometimes)
Which side chains can act to accept proteins
Arginine (12.5)
Lysine (10.79)
Tryosine (10.13)
Cysteine (8.33)
What is Chymotrypsin an example of ?
Acid-base catalysis and covalent catalysis
What is the active site amino acid for Chymotrypsin?
Histidine
How is the product freed in Chymotrypsin catalysis ?
Acid catalysis breaks the acyl-enzyme covalent bond by H from NH on Histidine binding to O to make OH on Serine- then breaking the bond between the O of serine and the C of the product
What are Metal Ion catalysis or co-enzymes used for ?
- Binding substrates in the proper orientation
- Mediating oxidation-reduction reactions
- Electrostatically stabilizing or shielding negative
What are Metalloenzymes? Name the Five common ones
Contain TIGHTLY bound mental ions Fe+2 Fe+3 Cu+2 Zn+2 Mn+2
What are Metal-activated enzymes? Name the 4 common ones
Contain loosely bound metal ions (remove 1 or 2 electrons to stabilize outer ring) Na+ K+ Mg+2 Ca+2
what are the cofactors used for activation transfer reactions?
Thiamine pyrophosphate (TPP) Coenzyme A (CoA) Biotin Pyridoxal phosphate (PLP)
How do activation transfer reactions utilize the coenzyme in the reaction?
The coenzyme will provide a specific type of catalysis and then form a covalent intermediate bond with the substrate for additional catalytic power
(after TPP has a Carbanion, it will be attracted to Pyruvates slightly positive Carbon and facilitate Decarboxylation of Pyruvate- forming Resonance ion)
What are the 6 general classifications of enzymatic reactions?
- Oxidoreductases
- Transferases
- Hydrolyases
- Lyases
- Isomerases
- Ligases
What is the difference between Oxidation-reduction cofactors and activation transfer cofactors?
The Oxidation-reduction cofactors do not form covalent bonds with the substrate
However, both are not good catalysts without participation from amino acid side chains on enzyme
What are the Cofactors used in oxidation-reduction reactions?
NAD+ (used as cofactor for Lactate Dehydrogenase)
FAD
Vitamin E
Vitamin C
What is an example of a Oxidoreductase reaction?
Dehydrogenases -
Lactate Dehydrogenase - catalyzes transfer of electrons from lactate to NAD+ to make NADH
What is a Transferase class enzyme?
catalyze transfer of a specific functional group between molecules
What are the 4 types of Transferase?
Kinases
Aminotransferases
Acyltransferases
Glycosyltransferases
What do Kinases do (transferase) ?
transfer phosphate groups (usually from ATP to another molecule)
-like hexokinase or glucokinase
What do Amniotransferases do?
Transfer amino groups that are important in amino acid metabolism (H3N)
What do Acyltransferases do?
Transfer fatty acyl groups
What do Glycotransferases do?
Transfer carbohydrate residues
What do Hydrolases enzymes do?
Hydrolysis reactions that cleave bonds by addition of a water molecule
Ex: Proteases (4 types)- Cleave peptide bonds
What are the 4 types of Proteases?
Cysteine Proteases ;Caspases
Aspartyl Proteases: BACE
Metalloproteases: Matrix metalloproteinases (MMPs)
Serine Proteases: Trypsin and Chymotrypsin
What do Cysteine proteases do ? (SH containing)
Caspases- key mediators of apoptosis
What do Aspartyl proteases do ? (BACE)
involved in the amyloid peptide generation of Alzheimers disease
What do Metalloproteases (Matrix metalloproteases MMPs) do?
family of enzymes that are responsible for the degradation of extracellular matrix components such as collagen, laminin, and proteoglycans
What do Serine proteases do (trypsin and chymotrypsin)?
Digestive enzymes in the small intestine
What do Lyases do ? what are the two example classes?
Enzymes involved in cleaving bonds by means other than hydrolysis or oxidation
Aldolases or Thiolases
What do Aldolases with the Lyases category do?
(Ex: Fructose diphosphate aldolase- involved in glycolysis in breaking Fructose 1,6 bisphosphate in half into 2 different 3C chains)
What do Thiolases do (in category of lyases)?
Involved in breakdown of fatty acids
What do Isomerases do?
Rearrangement of atoms of a molecule to create an isomer of the starting compound
-enzymes catalyzing this are called ISOMERASES
What is an example of an Isomerase?
Triose phosphate isomerase - moves around certain groups on the split 3C chains during glycolysis
What do Ligases do?
Synthesizing bonds between carbon atoms and :
Carbon
Nitrogen
Oxygen
Sulfur atoms
Coupled to cleavage of high energy phosphate of ATP (or other nucleotide)
What is an example of a Ligase?
Pyruvate carboxylase - key enzyme in gluconeogenesis , important ligase in metabolism
What is an example of a Ligase?
Pyruvate carboxylase - key enzyme in gluconeogenesis , important ligase in metabolism to create irreversible Oxloacetate from Pyruvate
Explain the function of Serine protease?
The serine functions next to histidine in the catalytic triad as an electron acceptor (nucleophilic)
-then the Oxyanion tetrahedral intermidate is formed with O on Serine and Serine and Glycine stabilize using hydrogen bonds
(Example: Chymotrypsin)
What is covalent catalysis ?
The substrate forms transient covalent bonds within the active site residue amino acids or coenzymes
(Example: Serine on the substrate after nucleophilic attack by Histidine H transfer from its NH)
What is the Michaelis-Menton Equation ?
Vi = Vmax[S] / Km + [S}
What does Vi stand for?
Initial Velocity
What does Vmax stand for?
Maximum Velocity
What does Km stand for ?
The enzyme concentration needed to reach 1/2 Vmax
reflection of the affinity of an enzyme for a substrate
What does a Small Km indicate ?
High affinity (low S concentration is needed to half saturate the enzyme) - Denominator
What does a large Km indicate?
Low affinity (high Substance concentration is needed to half saturate enzyme )
What are the assumptions of the Michaelis Menton equation?
That the substrate concentration is larger than enzyme
Enzyme is the rate limiting factor (not substrate)
Stead state assumption (no change over time)
Initial velocity is analyzed as the immediate time of mixing (allows ignoring the backward movement of reaction between E+S and ES)
What happens at Vmax?
The enzyme is 100% saturated with Substrate and the reaction will not occur any faster
What do Hexokinase and Glucokinase both do?
Accept Glucose as a substrate and phosphorylate it into Glucose - 6 phosphate
Where is Hexokinase found? Glucokinase?
Hexokinase is found in Brain
Glucokinase is found in Liver
What is the activity of Hexokinase?
very Hyperbolic graph
low Km (higher affinity), low Vmax (rapidly saturated)
Good so glucose is always avaliable in brain, and it can not be sequestered in the brain
What is the activity of Glucokinase?
slightly sigmoidal graph
-high Km(needs more substrate to reach 1/2 Vmax), high Vmax
Good for storage of excess energy (glucose- sequestered)
Effective for after a meal- can handle high concentrations- stores as glycogen
What 3 ways are enzymes regulated?
Inhibitors
Conformational changes
Regulation thru enzyme concentration
What are the 2 types of inhibitors ?
Reversible
Irreversible
Inhibitors are compounds that decrease the rate of reaction
What are the two types of Irreversible inhibitors ?
Covalent
Transitions state analogs (suicide inhibitors)
How does a covalent irreversible inhibitor function?
Covalent bonds form within the active site or with enzyme cofactors
What is an example of Irreversible inhibitors (covalent)?
Aspirin
- forms covalent modification to prostaglandin endoperodixe synthase
- then reduces the formation of prostaglandin, hence inflammation is reduced
How do irreversible inhibitors of the transition state analogs work?
Bind more tightly then the substrate would to active site
What is an example of the transition state analog for Irreversible inhibitors/
PCN- inhibits the bacterial enzyme glycopeptide transpeptidase
- PCN has a strained peptide bond- resembles the transition state more than the substrate does so it fits better into the active site (“step ahead”)
- Covalent attaches to the active site to shut down
What are the 3 types of Reversible inhibitors?
Competitive (bind active site)
Non-Competitive( bind in site alternative to active site)
Product inhibition
What is the equation for a lineweaver burk plot ?
1/V =Km/Vmax (1/[S]) + 1/Vmax
How do you find the X intercept for lineweaver burk plot ?
-(1/Km)
Km = 50% of Kmax or Kmax/2
How do you find the Y intercept on lineweaver burk plot?
1/Vmax
How do you find the slope of the lineweaver burk plot?
Km/Vmax
What is on the Y axis of lineweaver burk plot? X axis?
Y axis = 1/V
X axis = 1/[S]
Describe the lineweaver burk plot for the addition of a Competitive inhibitor?
Increase Km (X intercept moves to the right and Y moves down) - Slope is steeper No effect on Vmax
Describe the lineweaver burk plot for the addition of a non-competitive inhibitor?
No change on Km (X intercept stays the same)
Decrease in Vmax- affinity is same but less can literally bind (Y intercept moves up )
Slope steeper than with no inhibitor
Why does the Vmax on the Pure non-competitive inhibition decrease?
No level of increase in Substance concentration will outcompete the non-competitive inhibitor the way a competitive inhibitor can be outcompeted
Describe Reversible inhibition in relation to product inhibition ?
All products are reversible inhibitors of the enzymes that produce them
Ex: Hexokinase is inhibited by its product Glucose 6-P
What are 4 types of Conformation changes for enzyme regulation?
Cooperativity
allosteric activators
Protein-Protein
Cleavage
What is an example of a conformation change as enzyme regulation for cooperativity?
Hemoglobin- Oxygen binds with greater affinity after each conformation change induced by each O2 bond that occurs until it is saturated
What is an example of an allosteric activator (conformation changes)?
Acetyl CoA (lygase) allosteric activator of pyruvate carboxylase
What enzymes do phosphorylation ?
what do dephosphorylation ?
Kinases add phosphate groups
Phosphatases take away phosphate group
(both play important roles in regulation of metabolic enzymes)
What is the structure of the phosphate group that is added in phosphorylation?
4 Oxygen groups (one double bond, one bound to P and then substrate, and the other two have single bonds with negative charge)
What is an example of conformation changes (Phosphorylation) to regulate enzyme activity?
Ex: Muscle glycogen phophorylase B can be activated into two different forms that are two different conformational changes
- either by AMP (low energy state- makes glycogen)
- Kinase- adds phosphate group
What is an example of a conformation change for Protein-Protein interactions for enzyme regulation?
Monomeric G-proteins
Describe the function of how protein-protein interactions work for G-proteins?
- GTP binds and G-protein is active
- then both bind a target protein, and make it ACTIVE only when bound to G-protein
- Hydrolyse the GTP to GDP, then protein becomes inactive again and releases
- GDP needs to recharge into GTP for new cycle
What are one types of Conformation change by way of cleavage? Example?
Zymogens (require cleavage for activation)
Ex: Chymotrypsinogen
-stored in pancreatic cells
-converted to chymotrypsin via trypsin when needed
What is the importance thing about Zymogens?
Synthesized in an inactive form to ensure they do not inappropriately activate out of intended area (digestive enzymes)
What are the two ways to regulate enzymes through concentration?
Synthesis (Increase or decrease in transcription)
Degradation (Ubiquitination) “kiss of death”
What is allosterism ?
A change in activity and conformation of an enzyme resulting from the binding of a compound at a site on the enzyme other than active site
Ex: CO2 is allosteric inhibitor for O2 binding (changes shape and affinity)
