enzymes Flashcards
define enzymes
a catalyst that changes the rate of a reaction without being changed in the process
Properties of enzymes x4
- They are highly specific for which substrate they bind to and which product is formed
- There activity can be regulated and the activity can vary depending on the concentration of the substrate it is working on and other molecules
- Nearly all enzymes= globular proteins [ some RNA catalyst have been found though #ribozymes]
- Enzymes are made in a way where they are endogenous to the naturally occurring enzyme that may be found in the body [#look the same as the naturally occurring enzyme]
Definition of enzyme active site and it is usually a what on the enzyme
region of the enzyme that binds to the substrate and converts it into a product- usually a crevice [indentation]
What polarity does the active site typically have- does this have a benefit
hydrophobic - predominantly nonpolar- enhances binding
how is substrate bound to active site [forces]
multiple weak forces - [electrostatic, hydrophobic, hydrogen bonds] & some have reversible covalent bonds
once substrate and enzyme bind what is the product and what does this cause
enzyme-substrate complex is formed and the catalytically active A.A. which bind -> substrate and cause change its change into product - product is then released
once product is formed and released what happens
catalytic cycle begins again
how does the substrate bind to the Active site
- Interactions occur between the A.A. side groups which holds the substrate in place
- The A.A in active site actively participates in the enzymes catalysis
describe lock and key model
- Shape of substrate + enzymes active site fit together like a lock and key
- 2 shapes = rigid + fixed [#don’t change in shape]
They are Perfectly complement - don’t change in shape
- 2 shapes = rigid + fixed [#don’t change in shape]
describe induced fit model
- When the substrate binds it induces a conformational change
can some enzymes show different features of both molecules
yes- they have some complementarity and some complementary changes- give 2 scenarios
what determines an enzymes specifity
properties and spatial arrangement of A.A. residues determines enzyme specificity - i.e. which molecules can bind
- If there’s small A.A. - bulky substrate can fit in
- Large A.A. residues - mean small substrate can fit
how is specificity determined
- Determined by changes in a few A.A. in the active site
give 3 e.g.s of digestive enzymes and where are they made
- Trypsin
- Chymotrypsin
Elastase
Pancrease
- Chymotrypsin
what are the digestive enzymes called- why are they called this
serine proteases - They have serine residues [involved in catalysis]
- They are proteases as they catalyse the hydrolysis of peptide bonds in protein [i.e. substrate is also made of A.A.]
- The Enzymes cleave peptide bonds in their substrate [which are proteins] - cause slight conformational
- There specificity is based on nature of A.A in residues [complementary bind]
explain the enzyme specificity of chymotrypsin
- The enzyme cleaves on the carboxyl side of the bulk aromatic + hydrophobic substrate A.A.
- Enzyme’s A.A. has small chains which gives access to bulky side chains of substrates
enzyme specificity of trypsin
- Enzyme cleaves to carboxyl side of positively charged Lysine + Arginine
- This is due to the negatively charged asparagine it has in its active site - this interacts with arginine’s + lysine’s positive side chain [#substrates]
enzyme specificity of elastase
- Cleaves the carboxyl groups of small uncharged side chains of substrates
- Enzyme possess A.A. residues with large side chains so larger molecules cannot fit
- Alanine and glycine however can fit
what determines an enzymes specificity
- Shape + A.A. content at the active site determines which substrate can bind [#specicity]
- Each enzyme = usually substrate specific
what 2 stages are in a biochemical reaction
- Substrate is converted to transition state [halfway] molecule becomes distorted to an electronic conformation [energy require here is + because energy is being taken in]
- Transition state = converted to a product
energy hump [activation energy]
is a barrier to chemical reactions
whats an enzymes main function
Enzymes main function is to reduce the activation energy and increase rate of reactions sometimes by factor of 10^7 -> 10^14 this 10^14 means that because of the enzyme the reaction that would take a matter of years only takes 1 second
free energy charge is determined by what?
by free energy difference between starting substrate and product Overall reaction must have a negative free energy [meaning the energy of the reactants = greater than the energy of the products ] if its not negative the reaction will not occur
Enzyme doesn’t affect the overall change in energy rather just speeds it up [in terms of the ratio
define enzyme activity, specific activity, Kcat
I Unit is the amount of enzyme that converts 1 micromolar substrate per minute at 25 degrees Celsius at optimal pH
Specific activity = units of enzyme per mg of protein present
Turnover number [kcat]- maximum number of molecules of a substrate that an enzyme can convert to product per catalytic site per unit of time
who classifies enzymes
IUBMB - union biochemistry & molecular biology [enzyme commission
explain 4 digit numbering system for enzymes
1- Six classification numbers
2- Subclass [type of bound it acts upon]
3- A subclass[tells if cofactor is required + involved]
4- Is the serial number - it’s the order in which the enzyme was added to the list
e.g. alpha Amylase EC 3.2.1.1 - beta amylase EC 3.2.1.2
many enzymes require cofactors/coenzymes to carry out their functions
- Many enzymes require cofactors to carry out their particular reaction
- When a metal/coenzyme is covalently attached to the enzyme - this coenzyme/metal is called prosthetic group
- Complete catalytically-active enzyme with its prosthetic group = HOLOENZYME
- Protein part of enzyme without its prosthetic group = called apoenzyme
- Some coenzymes i.e. NAD+ = bound + released by enzyme during the catalytic cycle -i.e. function as co-substrates [part of the product]
- Many Coenzymes = derived from Vitamin precursors.
- An inadequate supply of these vitamins leads to deficiency disease
When a metal/coenzyme is covalently attached to the enzyme what is it called
a prosthetic group
complete catalytically-active enzyme with its prosthetic group is called what?
Holoenzyme
protein part of enzyme without its prosthetic group is called what?
apoenzyme
some coenzymes like NAD+ are ? during the catalytic cycle this makes them?
Bound and released - co-substrates
many coenzymes are derived from?
vitamin precursors
what 2 factors affect enzyme activity
Ph and temperature
describe pH optimum- describe effects of small changes and large changes- most enzymes are optimum at what ph- give exception
- Each enzyme has an optimum pH- [the rate of the reaction is optimum at this pH]
- Small changes cause small deviations due to change of ionisation [charge] of groups at the active site of the enzyme
- Large deviations cause the enzyme to be denatured due to weak noncovalent bonds being interfered [these are the bonds that maintain the enzymes 3D structure]
- Many enzymes are optimum at 7 pH because this is the bodies physiological pH
- Exception = digestive enzymes i.e. pepsin which optimum ph is near 2 as stomach is acidic so needs to have a low optimum to survive.
describe temperature optimum- small and large effects
- Small increase in temp increase the thermal energy of the substrates. This also lowers the energy activation and increase the rate of reaction
- Large increase in temp bonds [weak non-covalent reactions that hold enzyme in it’s 3D form = broken - protein becomes denatured
- Most mammalian enzymes = 37 degrees - why? Because its physiological temperature
Enzyme inhibitors - what are they and what do they do and give 2 examples of them
- Inhibitors are molecules that work directly on enzymes -> lower its catalytic rate
- e.g. normal body metabolites [inhibits a particular enzyme in normal metabolic control
- foreign substances i.e. drugs/toxins effect enzyme inhibition can be therapeutic or lethal
enzyme inhibition can be x or y
inhibition can be reversible or irreversible
reversible inhibition of enzymes can be? x3
- competitive [blocks active site so substrate won’t go in]
- Non-competitive [change occurs at active site - substrate cant go in
- Uncompetitive
Isoenzymes are defined as
different forms of an enzyme that catalyse the same reaction
These isoenzymes exhibit ? and derived from?
exhibit different physical + kinetic properties i.e. optimum pH
- They are also derived from different genes which occur in different tissues of the body
give an example of an isoenzyme and what is does
catalyzes reversible reaction conversion of pyruvate into lactate in the presence of the coenzyme NADH. LDH = tetramer made up of 2 different types of subunits [H & M]
LDH is a tetramer made of 2 different subunits which combine to form different subunits give 5 possible isoenzymes
- H4 [predominantly in heart + red blood cells]
- M4 [predominantly in skeletal muscle + liver]
- H3M [predominantly in heart + RBC]
- HM3 [predominantly in liver + skeletal muscle]
- H2M2 - in brain and kidney
- M units = found in skeletal muscle + liver
- H subunit - found in heart
what are the allosteric enzymes
enzymes that change their shape, or
conformation, upon binding of an effector molecule
the biological activity of allosteric enzymes = effected by what?
by altering
the conformation of its tertiary structure
allosteric enzymes tend to have how much subunits
several subunits
in some cases the regulatory site which binds the effector molecule and the x are on ? [allosteric enzymes]
active site - are on separate subunits
define an allosteric effector
is a substance that modifies the behaviour
of allosteric enzymes; it may be an allosteric inhibitor or
allosteric activator.
what does an allosteric activator
increases the rate of enzyme activity,
while an allosteric inhibitor decreases the rate of enzyme
activity.
explain competitive inhibition
- its reversible and - reduces the amount of
enzyme available for
substrate binding - may be overcome by increasing substrate - molecule goes in there and attaches to active site
non-competitive inhibition-reversible
this inhibitor doesn’t bind to the catalytic site rather binds somewhere else to change enzymes shape - binds -> allosteric site or regulatory site
distinguish between allosteric inhibition and allosteric activation
both types have molecules that bind @ sites other than active site and one activates the substrate to bind to the same enzyme whilst the other prevents the substrate binding to the same enzyme
un-competitive inhibition
is also reversible - intermediate substrate is formed and inhibitor binds to this enzyme substrate - no product is formed
some allosteric activator are the s- itself
these types are said to exhibit what charge cooperativity
define this charge cooperativity
describe the binding of substrate molecules
substrate itself- positive cooperativity
a positive charge c- is when binding to one subunit facilitates binding of substrate to another subunit
the first substrate has difficulty binding-> enzyme as all subunits = in conformation with a low affinity for the substrate - [T-conformation i.e. inactive] when the 1st substrate binds @ least 1 adjacent subunit is changed to the high-affinity conformation [R state- relaxed]
when does the activator bind to it’s activator site
when the enzyme is in the R configuration- allows substrate to bind
when is the inhibitor binding site of an enzyme open- what does this do the the enzyme
when enzyme is in the T [inactive state] -keeps active site closed/ less accessible- enzyme = inactive
give an example of an allosteric inhibitor
can be the product of the enzymatic reaction- if there’s too much product it will bind -> allosteric inhibitor site and enzymatic reaction = stopped #negative feedback inhibition
give 3 examples of irreversible inhibitors
- transition-state analogues
- covalent inhibitors
- heavy metal inhibitors
what are transition-state analogues
- Compounds that resemble the substrate
when its in the transition-state - Bind to active site and keep the reaction at
that point - Reaction cannot progress and free up active
site for new substrate molecule
E.G– Penicillin inhibits the enzyme transpeptidase - Transpeptidase – catalyses the formation of the peptide cross links in
peptidoglycan - Penicillin – has beta lactam ring – resembles the special amino acid found in the peptide links
- Forms irreversible bond with catalytic serine in the transpeptidase active site
what are covalent inhibitors
Compounds that react irreversibly with
amino acids in the active site e.g. aspirin* Diisopropyl phosphofluoridate (DFP) is a neurotoxin
* Forms covalent intermediate in Acetylcholinesterase active site
* Acetylcholinesterase = enzyme that degrades the
neurotransmitter acetylcholine (sends messages in the brain) that’s why its called neurotoxin
what are heavy metal inhibitors is it specific? give examples
Heavy metals that bind to important amino acids in the active site and prevent them from taking part in substrate binding or
catalysis- usually non-specific- mercury, lead, aluminium and Fe
Lead replaces important metals that act as an enzyme co-factor
* Replaces Ca2+ ions in calmodulin and protein kinase C
give 4 examples of reversible inhibitors
- competitive
- non-competitive
- uncompetitive
- allosteric inhibitors
irreversible inhibitors examples x3
- transition-state analogues
-covalent inhibitors
-heavy metals
