Enzymes Flashcards
True or false: enzymes are catalysts, that are not consumed in processes.
True
Nearly all enzymes are _________ __________, although a few catalytically-active RNA molecules (ribozymes) have been identified.
Globular proteins
What is an enzyme’s active site?
The region of the enzyme that binds into the substrate, and converts it into product
The active site is often a cleft or crevice on the surface of the enzyme, that forms a predominately non-polar, or ______________, environment, to enhance substrate binding.
Hydrophobic
List four forces which bind the substrate to the active site.
Electrostatic interactions, hydrogen bonds, and hydrophobic forces, and in some instances, reversible covalent bonds
What adjective describes the molecule that the enzyme substrate complex creates?
Bound
Catalytically-active _______ ______ residues within the active site act on the substrate molecule, to transform it into product, which is then released.
Amino acid
At a molecular level, the __________ fits into the active site.
Substrate
Specific interactions with the amino acid side groups at the _______ ______ hold components in place.
Active site
Describe the enzymatic ‘lock-and-key’ model.
The lock-and-key model elucidates the theory that the shape of the substrate and the active site are rigid, perfectly complementary, and fit together exactly, when in the right alignment
Describe the enzymatic ‘induced fit’ model.
In the induced-fit model, the binding of the substrate induces a conformational change in the active site of the enzyme
True or false: some enzymes show features of both models, with some complementarity.
True
To what does ‘enzyme specificity’ refer?
How the properties and spatial arrangement of the amino acid residues forming the active site determine which molecules can bind and become substrates
List three enzymes in which enzyme specificity is observed.
Trypsin, chymotrypsin, and elastase
To what family do trypsin, chymotrypsin, and elastase belong?
Serine proteases
Explain the etymology of ‘serine proteases’.
‘Serine’ comes from the serine residue in the active site that plays a critical role in catalysis, and ‘protease’, from the catalysis of the hydrolysis of peptide bonds in proteins
Describe the enzymatic action of chymotrypsin.
Chymotrypsin cleaves on the carboxyl side of bulky, aromatic, and hydrophobic amino acid residues. Amino acid residues with small side chains in its active site allow for access to bulky side chains of substrates
Describe the action of trypsin.
Cleaves on the carboxyl side of positively charges lysine or arginine residues, and has a negatively charged Asp residue in its active site, to react with Lys and Arg
Describe the action of elastase.
Cleaves on the carboxyl side of amino acid residues with small, uncharged side chains. It has relatively large uncharged side chains of Val and Thr protruding into its active site, preventing access of all but the small side chains of Gly and Ala
What is ‘energy of activation’?
The changes that take place during the course of a particular process
In the first stage, ___ must be converted to the S† transition state.
S
In the second stage, S† is converted to ___.
P
True or false: the S† is at a lower free energy level than S.
False
The free energy change for S to S† is __________, and termed the ‘energy of activation’.
Positive
The _________ ______ constitutes a barrier to chemical reactions occurring.
Energy hump
______ __________ ___________ between the starting substrate and final product impacts the reaction.
Free energy difference
What is the enzymatic ‘turnover number’?
The maximum number of molecules of substrate that an enzyme can convert to product, per catalytic site, per unit time (kcat)
In enzyme nomenclature, the first number has _____ classification numbers.
Six
The second and third numbers in enzyme naming are __________.
Subclasses
True or false: in enzyme naming, the fourth digit is a serial number.
True
Many enzymes require the presence of small, nonprotein units, called __________ to carry out their particular reaction.
Cofactors
Cofactors may be either one or more _________ ions, such as Zn2+ or Fe2+, or a complex organic molecule called a coenzyme.
Inorganic
A metal or coenzyme that is covalently attached to the enzyme is called a ____________ ________.
Prosthetic group
A complete catalytically-active enzyme together with its prosthetic group is called a ______________.
Holoenzyme
The _________ part of the enzyme on its own, without its prosthetic group, is called an apoenzyme.
Protein
Some coenzymes, such as NAD+, are bound and released by the enzyme during its catalytic cycle and so function as ____-___________.
Co-substrates
Many coenzymes are derived from _________ precursors.
Vitamin
Each enzyme has an __________ pH.
Optimum
Rate of reaction is optimum at at an enzyme’s special pH, and ________ is at its prime.
Efficacy
Small deviations from pH optimum cause decreased activity, due to changes in the ____________ of groups at the active site of the enzyme.
Ionisation
Larger deviations from pH optimum causes _______________ of the enzyme itself, due to interference with the many weak noncovalent bonds maintaining its 3D structure.
Denaturation
Many enzymes have an optimum pH around ____________ pH.
Physiological
What is pepsin’s optimum pH?
~2
Small rises in temperature increase the thermal energy of substrate molecules, ____________ the energy of activation, increasing rate reaction.
Lowering
True or false: large rises in temperature denature the enzyme.
True
What are enzyme inhibitors?
Molecules that are capable of interfering with the activity of an individual enzyme
A molecule which acts directly on an enzyme to lower its ___________ ______ is an inhibitor.
Catalytic rate
List two examples of inhibitors.
Normal body metabolites, which inhibit a particular enzyme as part of the normal metabolic control of a reaction, and foreign substances, such as drugs or toxins - the effect of enzyme inhibition could be either therapeutic or, at the other extreme, lethal
True or false: inhibition is always reversible.
False
__________ inhibition can be either competitive, noncompetitive, or uncompetitive.
Reversible
What are isoenzymes?
Different forms of an enzyme which catalyse the same reaction
_____________ exhibit different physical or kinetic properties, e.g., isoelectric point, pH optimum, substrate affinity, or effect of inhibitors.
Isoenzymes
Isoenzymes are usually derived from different _______, and often occur in different tissues of the body.
Genes
Cytochrome P450 is a superfamily of membrane-bound ____________ isozymes.
Haemoprotein
CYP predominately occupy the ________, intestines, and kidneys.
Liver
Six of the 57 P450 isoenzymes are responsible for 90% of ______ ____________.
Drug metabolism
What does lactate dehydrogenase do?
Catalyses the reversible conversion of pyruvate into lactate in the presence of the coenzyme NADH
Lactate dehydrogenase is a ___________ of two different types of subunits, called H and M, with small differences in amino acid sequence between subunits.
Tetramer
List the five isoenzymes of lactate dehydrogenase.
H4, H3M, H2M2, HM3, and M4
____ subunits of LDH found mostly in skeletal muscle and liver.
M
H subunits of LDH predominate in the ______.
Heart
H4 and _______ LDH isoenzymes are found predominantly in the heart and red blood cells.
H3M
The H2M2 __________ ________________ isoenzyme is found predominantly in the brain and kidney.
Lactate dehydrogenase
HM3 and ______ LDH isoenzymes are found predominantly in the liver and skeletal muscle.
M4
List three reasons as to why enzymes are important.
Causes of disease, in the event of mutated enzymes
Food processing
Drug manufacture
_____________ inhibitors reduce the amount of enzyme available for substrate binding; they may be overcome by increasing the substrate.
Competitive
Non-competitive inhibitors do not bind to the ___________ site; they bind elsewhere (an allosteric, or regulatory, site), distorting the enzyme’s shape.
Catalytic
_______________ inhibitors have irreversible effects; they can only bind to the enzyme substrate complex, and no product is formed.
Uncompetitive
____________ enzymes change their shape or conformation, upon binding with an effector molecule.
Allosteric
The biological activity of allosteric enzymes is impacted by altering the __________ structure.
Tertiary
True or false: allosteric enzymes tend to only have one or two subunits.
False
In some cases, in the context of allosteric enzymes, the ___________ site, which binds the effector molecule, and the active site are on separate subunits.
Regulatory
An allosteric ___________ may be an inhibitor (impeding) or activator (promoting), and modifies the behaviour of allosteric enzyme.
Effector
If there is too much product present, the __________ binds to the allosteric inhibitor site, and the enzyme reaction is stopped.
Product
Sometimes, the allosteric activator is the ___________ itself.
Substrate
Enzymes are said to exhibit __________ ____-____________ (the binding of substrate to one subunit facilitates the binding of substrate to another subunit).
Positive co-operativity
The first substrate molecule has difficulty in binding to the enzyme, as it is in the _____ ‘T’ conformation.
Taut
The relaxed ‘R’ state follows, when the substrate changes its own ___________ to bind.
Subunit
Describe irreversible inhibitors’ transition state analogue.
Compounds resemble the substrate when in its transition state
Binds to the active site, and maintains the reaction
Reaction cannot proceed; for instance, penicillin inhibits transpeptidase
Transpeptidase catalyses the formation of peptide crosslinks in peptidoglycan, and penicillin, with its beta-lactam ring, binds irreversibly with serine at the active site
__________ inhibitors react irreversibly with amino acids in the active site, such as aspirin.
Covalent
___________ ________ __________ bind to important amino acids in the active site, and prevent them from taking part in substrate binding and catalysis.
Heavy metal inhibitors
Lead and mercury can bind tightly, but are _____-__________.
Non-specific
Mercury often binds to _______ groups.
Thiol
Lead replaces important metals that act as __________ _____-__________, and replaces Ca2+ ions in calmodulin and protein kinase C.
Enzyme co-factors
DFP is a neurotoxin, that forms ___________ intermediate with acetylcholinesterase active site. This causes degradation to neurotransmitters.
Covalent
____________ inhibitors reduce the amount of enzyme available for substrate binding, and may be overcome by increasing the substrate.
Competitive
____________ inhibitors cause no product to be formed; the inhibitor binds the enzyme-substrate complex, but increasing the substrate has no effect.
Uncompetitive
Binding of the inhibitor affects both the Km and the _______ _____.
True kcat
Non-competitive inhibitors bind to sites involved in _________ and catalysis.
Binding
In the context of non-competitive inhibitors, inhibitors bind to the _________ site, the Km does not change, and the apparent Km and kcat change.
Allosteric
In general, uncompetitive drugs give the best inhibition, but ___________ inhibitors tent to be more specific.
Competitive
Binding affinity is determined by ______.
Km
The _________ _______ that an enzyme has for a drug is most important; the lower the dosage, the better.
Binding affinity
The initial rate of a reaction, ____, is measured at the start of the reaction.
Vo
Vo increases almost linearly with an increase in substrate concentration initially, until the __________ _______ (Vmax), is achieved.
Saturation point
Km (the Michaelis-Menten constant) is the concentration of substrate required for the enzyme to reach _______ of its Vmax, and is always expressed as mM.
Half
The lower the Km, the tighter the substrate binds to the enzyme, and a low Km means the enzyme has a high ___________ for substrate.
Affinity
A _______ Vmax means the reaction will occur quickly.
High
True or false: each enzyme has a unique Km and Vmax, such that specific enzymes can be chosen to suit a situation.
True
_________ inhibits cyclooxygenase, stops prostaglandin synthesis, and inhibits pain.
Aspirin
_________ inhibit HMG-CoA reductase in the liver, which controls heart disease.
Statins
Taxol inhibits ___________ _______, to prevent mitosis, and inhibit cancer.
Microtubule proteins
___-____ _____________ is useful in looking at the interaction between inhibitor and enzyme active site/site of action of the inhibitor, and aids the design of new classes of pharmaceuticals.
3-D modelling
The standard way of testing if an inhibitor compound is present is to examine the enzyme reaction, in the presence of a ________ ____________. An inhibitor should alter enzyme kinetics. Other factors, such as pH and heat, can also affect the rates of an enzyme reaction.
Potential inhibitor
Michaelis-Menten plots are often transposed into _____________-______ plots, whereby the reciprocal of V0 versus the reciprocal of [S] (that is, 1/V0 and 1/[S]), is plotted.
Lineweaver-Burk
_______ is the turnover number of an enzyme; this refers to the maximal number of substrate molecules converted to product, per active site, per unit time.
kcat
Apparent Km, or Kmapp, refers to the ___________.
Inhibitor
E + S <=> ES <=> E + P, where K+1 and K-1 are above and below the left arrows, and K+2 and _____ are above and below the right arrows.
K-2
What is lysozyme?
An antibacterial enzyme, forming part of the innate immune system
List the two major forms of lysozyme.
Human, and hen’s egg white
In Gram-negative bacteria, the _________________ acts as a non-competitive inhibitor, by highly-favoured binding with lysozyme.
Lipopolysaccharide
Lysozyme, like all enzymes, has a unique substrate, namely _________________.
Peptidoglycan
_____________ catalyse the hydrolysis or hydrolytic cleavage of chemical bonds, by the reaction A - B + H2O -> A - OH + B - H.
Hydrolases
Lysozyme reaction is the hydrolysis of the β(1-4) glycosidic bond between N-acetylglucosamine (NAG) and _______.
NAM
129 amino acids are contained within ______________, with one polypeptide chain.
Lysozyme
Lysozyme catalyses the hydrolysis of the β(1-4) ___________ _______ between NAM and NAG.
Glycosidic links
List the components of the peptidoglycan wall.
Polysaccharide backbone, composed of alternating NAG, NAM, sugar units joined by β(1-4) glycosidic bond, etc.
Lysozyme has two domains, with two ____________ bonds in each domain.
Disulphide
By constructing molecular models of lysozyme, it was found that the cleft could be completely occupied by six ________ ____________ (labelled as A-F), with hydrogen bonding between them.
Sugar residues
Different lengths of NAG NAM ____________ 2, 3, 4, 5, 6, 7, 8 have been tested.
Polymers
True or false: for lysozyme, polymers 2-5 were poor substrates, but polymers 6 and upwards were effective.
True
In lysozyme, it was deduced that cleavage occurs between the NAM (D residue) and NAG (E residue), and that six residues were ___________ at the target site.
Optimal
Modelling further showed that two ___________ residues are near hydrolysed bonds in lysozyme.
Acidic
At optimum pH for lysozyme (pH ___), it would deprotonate some substances, and protonate others.
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