Enzyme: Properties, functions, classification

What are enzymes? Are they consumed during the reaction?
Enzymes are a class of proteins that act as catalysts and increase the rates of biochemical reactions. They are not consumed during the reaction.

Describe the general properties of the structure of an enzyme.
Like all proteins, enzymes are linear chains of amino acids that fold to produce a three-dimensional structure. The sequence of the amino acids specifies the structure which in turn determines the catalytic activity of the enzyme.
Enzymes have an Active site. The active site consist of the catalytic site and the binding sites. The catalytic site is located next to one or more binding sites where residues orient the substrates.
The structure of the catalytic site is essential for activity. If you alter the structure, you alter the function.

What are the factors that will influence enzyme activity?
Factors that influence enzyme activity include:
Concentrations of reactants (substrates, coenzymes, cofactors), products, and enzyme
Temperature
pH
and the presence of Activators, inhibitors, and inactivators

How does increasing the concentration of substrate affect the enzyme reaction rate?
Increasing the substrate concentration will increase the reaction rate until a certain point. This is because initially there is more substrate colliding with enzyme molecules, resulting in more product. However, at a certain concentration, the enzyme molecules will become saturated and the substrate concentration will no longer be the limiting factor.

How does increasing the concentration of enzyme affect the reaction rate?
Like increase the substrate concentration, increasing the [enzyme] will increase the reaction rate until a saturation point in which the enzyme is longer the limiting factor.

How does temperature influence enzyme activity?
An increase in temperature will increase activity to a point at which the enzyme becomes denatured. At which point the enzyme activity declines

How does pH influence enzyme activity.
pH dependency varies among enzymes. However, like temperature, there exists a denaturation point, and an optimal pH at which the enzyme activity is the highest leading to a bell shape curve on a enzyme activity/pH plot.

What are activators, inhibitors and inactivators and how do they influence enzyme activity?
Enzyme activators are molecules that bind to enzymes and increase their activity. They are the opposite of enzyme inhibitors, that bind to enzymes and inhibit their activity. These molecules are often involved in the allosteric regulation of enzymes.

How are enzymes classified?
Enzymes are classified based on the type of reactions they catalyze.

What are the different enzyme classes?

1. Oxido-reductases (oxidation-Reduction rxn that usually require coenzymes)
2. Transferases (Group Transfers)
3. Hydrolases (Cleavage of Bonds w/ addition of H20)
4. Lyases (Breaking of Bonds w/out addition of H20)
5. Isomerases (Racemization of optical or geometric isomers)
6. Ligases (Bond Formation with energy supplied by some high energy compound like ATP)

What are coenzymes? What are prosthetic groups?
A coenzyme is a nonprotein compound that is necessary for the functioning of an enzyme and are considered reversible. A prosthetic group is a nonprotein group forming part of or combined with a protein and is considered a irreversibly bound coenzyme

What are examples of coenzymes with Oxido-reductases?
NAD+ / NADH + H+ (lactate/alcohol dehydrogenase)
NADP+ / NADPH + H+ (cytochrome p450 reductase)
FAD / FADH2 (succinate dehydrogenase)
Cu2+ / Cu+ (cytochrome c oxidase)

What are examples of coenzymes with Ligases?
ATP; Biotin are used by Pyruvate Carboxylase

What are examples of coenzymes with Transferases?
ATP / ADP used by numerous kinases
Pyridoxal phosphate used by various aminotransferases

Where are enzymes distributed?
Enzymes are distributed within cells and within tissues depending on the type of reactions required. For examples, Hydrolases are found in lysosomes, Peroxidases are found in peroxisomes and DNA polymerases are found in the nuclei.

How do we get different forms of enzymes that catalyze the same reaction?
Many enzymes are synthesized by pro-enzymes (called zymogens), in a way that the actual catalysis is deferred until something else happens to that sequences. For example, with pepsinogen, part of the amino terminus is stuck over the catalytic site except at low pH revealing the active site leading to Pepsin.
Every pepsin formed may have a slightly different terminus.

Something must occur

What are Isozymes? What is an example?
Isozymes are multiple forms of reaction-specific enzymes with genetically determined differences in polypeptide amino acid sequences. They have identical functions but different structures. An example is Lactate Dehydrogenases (LDH)

What are the properties of Lactate Dehydrogenase (LDH)
Lactate dehydrogenase is an enzyme found in nearly all living cells and is composed of four subunits (tetramer). It catalyzes the conversion of lactate to pyruvic acid and back. The two most common subunits are the LDH-M and LDH-H protein, encoded by the LDHA and LDHB genes, respectively. These two subunits can form five possible tetramers (isoenzymes): 4H, 4M, and the three mixed tetramers (3H1M, 2H2M, 1H3M). These five isoforms are enzymatically similar but show different tissue distribution: The major isoenzymes of skeletal muscle and liver, M4, has four muscle (M) subunits, while H4 is the main isoenzymes for heart muscle in most species, containing four heart (H) subunits.

What are the 5 isozymes of Lactate Dehydrogenase and where are they found?
LDH-1 (H4)—in the heart and in RBC (red blood cells), as well as the brain.
LDH-2 (H3M)—in the reticuloendothelial system
LDH-3 (H2M2)—Brain and kidneys
LDH-4 (HM3)—in the kidneys, placenta, and pancreas
LDH-5 (M4)—in the liver and skeletal muscle

How can Lactate Dehydrogenase be used as a cardiac marker?
LDH-1 isozyme is normally found in the heart muscle and LDH-2 is found predominantly in blood serum. A high LDH-1 level to LDH-2 suggest a Myocardial Infarction. LDH levels are also high in tissue breakdown or hemolysis

What are Interconvertable forms of enzymes? What are some examples?
Interconvertable enzymatic forms refers to the phosphorylation or dephosphorylation of the enzyme results in more than one form. Two examples are:
Glycogen phosphorylase b ⇄ Glycogen phosphorylase a’
Glycogen synthase a (I)’ ⇄ Glycogen synthase b (D)
‘=more active form

1.An increase in the concentrations of _ can increase the rate of enzyme catalyzed reactions.
A. Enzymes
B. Substrates
C. Inhibitors
D. Choices A and B only are both correct.
E. Choices A, B, and C are all correct.

1. D Increases in both enzyme concentration (A) and substrate concentration (B) can increase reaction rates. Inhibitors (C) would decrease rates.
2. Which one of the following is most associated with “denaturation” of enzymes?
   A. pH
   B. cofactors
   C. inhibitors
   D. reaction products
   E. prosthetic groups
3. A The pH (A) can be associated with “denaturation” of enzymes (as well as other proteins). Choices B-E influence enzyme function, but would not be directly associated with “denaturation”.
4. Kinases are classified as _.
   A. lyases
   B. isomerases
   C. hydrolases
   D. transferases
   E. oxidoreductases
5. D Kinases are classified as transferases.
6. Proteases (e.g. pepsin) are classified as _.
   A. transferases
   B. isomerases
   C. hydrolases
   D. lyases
   E. ligases
7. C Proteases are classified as hydrolases (as are other digestive enzymes).
8. Dehydrogenases are classified as __.
   A. lyases
   B. ligases
   C. hydrolases
   D. transferases
   E. oxidoreductase
9. E Dehydrogenases are classified as oxidoreductases.
10. NAD+ is a coenzyme commonly associated with __.
    A. oxidoreductases
    B. transferases
    C. isomerases
    D. hydrolases
    E. ligases
11. A NAD+ is commonly associated with oxidoreductases.
12. ATP is a coenzyme commonly associated with __.
    A. hydrolases
    B. isomerases
    C. oxidoreductases
    D. transferases
    E. lyases
13. D ATP is a coenzyme commonly associated with (D) transferases (e.g. kinases) and with ligases (where the energy required to form covalent bonds is provided by ATP).
14. Lysozomes contain predominantly __.
    A. oxidoreductases
    B. transferases
    C. isomerases
    D. hydrolases
    E. ligases
15. D Lysozomes contain predominately acid hydrolases.
16. Isozymes are classified as different forms of enzymes resulting from _.
    A. phosphorylation
    B. partial proteolysis
    C. covalent modifications
    D. genetically determined differences in catalysis
    E. genetically determined differences in primary structure
17. E. Isozymes have genetically determined differences in their primary structures (i.e. amino acid sequences). They catalyze the same biochemical reactions; therefore catalysis would be very similar, thus (D) is false. Both phosphorylation (A) and covalent modifications (C) might be associated with interconvertable enzymes, and both partial proteolysis (B) and covalent modifications (C) could be associated with proenzyme (zymogen) partial proteolysis.
18. Which of the following lactate dehydrogenase isozymes would increase in serum as a result of acute hepatitis?
19. LDH2 (H3M)
20. LDH3 (H2M2)
21. LDH4 (HM3)
22. LDH5 (M4)
    A. Choices 1, 2 and 3 only are correct
    B. Choices 1 and 3 only are correct
    C. Choices 2 and 4 only are correct
    D. Choice 4 only is correct
    E. All choices are correct.
23. E (All choices) Liver tissue contains LDH M subunits, therefore the LDH
    isozymes in the serum with M subunits would increase following acute hepatitis.
24. Which of the following isozymes would increase in serum as a result of hemolysis?
25. LDH2 (H3M)
26. LDH3 (H2M2)
27. LDH4 (HM3)
28. LDH5 (M4)
    A. Choices 1, 2 and 3 only are correct
    B. Choices 1 and 3 only are correct
    C. Choices 2 and 4 only are correct
    D. Choice 4 only is correct
    E. All choices are correct.
29. A (Choices 1, 2 and 3) Red blood cells contain LDH H subunits, therefore
    the LDH isozymes in the serum with H subunits would increase following hemolysis.
30. Enzymes that have more than one form based on phosphorylation or dephosphorylation are classified as interconvertable enzymes.
    A. True
    B. False
31. A (True) Phosphorylation and dephosphorylation of interconvertable enzymes (and other proteins) is very important in regulating cell function.

An enzyme is a protein that facilitates a cellular metabolic process inside an organism or a cell by lowering the activation energy

Activation energy is the minimum energy that is required for a reaction to proceed.

Example: Lactate dehydrogenase (LDH or LD) which converts lactate to pyruvate and back.

Coenzyme

A coenzyme is a substance aids the function of the enzyme, acting as a helper molecule for a biochemical reaction. Example: Nicotinamide adenine dinucleotide (NAD) in lactate dehydrogenase.

Apoenzyme

Apoenzyme is the protein component that is considered incomplete enzyme without a coenzyme. 

Cofactor

Cofactor serves the same function as coenzymes as they regulate, control, and adjust enzymatic activities. The main difference between the two is that cofactors are inorganic while coenzymes are organic. An example of a cofactor is Mg2+ which is required by hexokinase in the phosphoryl group transfer.

Holoenzyme

When an enzyme lacks the needed cofactor, it is called apoenzyme. When the enzyme has the required cofactor, it is known as the holoenzyme. Examples of holoenzymes include DNA polymerase which is composed of several protein subunits.

Properties of enzymes.

Catalytic activity

A very small amount of enzyme is enough to convert a large number of substrates and the enzyme remains unchanged.

Specificity

Enzymes are very specific in their action. They catalyze specific types of reactions and acts on a particular substrate only. Enzymes show different types of specificity:

• Bond specificity – E.g. Lipase is specific for ester bond in lipid
• Group specificity
• Substrate specificity – E.g. Arginase acts only on arginine
• Optical specificity – E.g. L amino acids oxidase acts on L-amino acids
• Co-factor specificity
• Geometric specificity

Reversibility

Most enzyme-catalyzed reactions are reversible while others are irreversible.

Sensitivity to heat and temperature

Enzymes are very sensitive to heat and temperature. Low temperatures deactivate enzymes, while high temperature increases enzymatic activity up to the optimum temperature, after which a further increase causes deactivation of the enzyme.

Sensitivity to pH

Enzymes are very specific to changes in pH. Some enzymes are active in acidic conditions, others in basic, and others in neutral pH.  

Functions of enzymes

Degradation of macromolecules into smaller fragments. An example is the degradation of proteins by protease into amino acids.

Signal transduction through a series of molecular events. An example is protein kinases in phosphorylation reactions.

Energy generation to power biological activities. An example is enzymes in glycolysis such as hexokinase.

Acting in ion pumps to move ions across the plasma membrane against the concentration gradient in active transport. An example is ATPases.

Clearance of foreign substances from the external environment or those produced by the body and not useful to the body. Examples include the cytochrome P-450 enzymes that control the rate at which many drugs are metabolized.

Classification of enzymes

• Oxidoreductases – transfer electrons from one molecule to another
• Transferases – catalyze the transport of functional groups from one molecule to another
• Hydrolases – catalyze hydrolysis, breaking bonds by water

Other types:

• Lyases
• Isomerases
• Ligases

Active site

• The active site is the part of an enzyme where the substrate must bind for the products to be formed. It is a groove or pocket region of the protein formed by the folding of the protein.
• The substrates must combine with the enzyme in a specific way for the reaction to proceed and form the products.
• The enzyme attracts substrates to its active site, catalyzing a chemical reaction and then allowing products to dissociate.
• The combination formed by an enzyme and substrate is called the enzyme-substrate complex.

Types of enzyme inhibition

Competitive inhibition – This occurs when a molecule that resembles the substrates of the enzyme, binds on the active site and prevents further binding of the substrate.

Non-competitive inhibition – This occurs when an inhibitor that does not resemble the substrate binds on a different site on the enzyme other than the active site.

Uncompetitive inhibition – This occurs when an inhibitor binds on the enzyme-substrate complex preventing formation and dissociation of products.

Suicide inhibition – This is when an inhibitor binds irreversibly to the enzyme through covalent bonding.