Enzymes

are proteins that catalyze the thousands of different chemical reactions that constitute our metabolism large molecules made from 20 amino acids short lifetime speed up chemical reactions by lowering the energy barrier to a reaction ---called the energy of activationso that the reaction takes place at the low temperature of an organism (37 C for a human). are essential to reduce the energy barrier or energy of activation

Substrate
The molecule (or molecules) that an enzyme acts on which is converted into a product or products. Example: When we digest dietary protein (the substrate) with proteindigesting enzymes in our small intestine, we produce amino acids and small peptides as products.

Active Sites
Represents the both the binding and catalytic domains of the enzyme protein Are typically clefts in the threedimensional enzyme structure, where amino acid residues from close or distant parts of the molecule can act on the substrate.

Binding Site

The enzyme binds the substrate

Catalytic Site

The substrate is converted to product

Types of Reactions

Irreversible Reactions are typically described as non-equilibrium reactions. Reversible Reactions- are commonly called equilibrium reactions, and the two terms mean the same thing.

Enzyme Catalyzed Reactions

PROVISION OF REACTIVE GROUPS BY COFACTORS

Cofactors
Any substance that needs to be present in addition to an enzyme to catalyze a certain reaction Other reactive groups not available on amino acids

Types of Cofactors

Coenzymes- organic molecules like vitamins and minerals Metal Ions- such Zn, Mg, or Mn Apoenzyme- the polypeptide part of the enzyme which needs cofactor to be activated Holoenzyme- an active compound formed by combination of a coenzyme and an apoenzyme

Prosthetic Group
a cofactor tightly bound to the enzyme at all

times a non protein component of an enzyme that is involved in the mechanism of the reaction

Example: Heme

bound tightly to hemoglobin and myoglobin

It is the deep red, non protein, iron-

containing component of hemoglobin that carries oxygen

B-Vitamins
There are 8 B vitamins we need to form the basic components for coenzymes Inadequate intake leads to deficiency diseases Leads to insufficient catalytic power of enzymes because they lack their coenzymes

Minerals

Zinc is an especially useful mineral in that it is an essential component of more than 300 different proteins, including enzymes for synthesizing RNA ( ribonucleic acid) and DNA, pancreatic digestive enzymes, and enzymes involved in carbohydrate, fat, protein and alcohol metabolism. Magnesium is associated with the energy rich ATP molecule, so it is involved in virtually all aspects of our metabolism.

Calcium act in signal transduction Sodium and Potassium are critical to the chemical composition of intracellular and extracellular fluids.

SP

S (substrate) is converted to P (product) catalyzed by enzyme E. P is a substrate of the reverse action. (equilibrium reaction) Goal is to measure the rate of this reaction to in the direction SP.

P will be formed, as S is mixed with E. As the concentration of P increases and that of S decreases, the reverse reaction will take place, becoming more important as the concentration of P increases. (The reaction rate in the direction toward P decreases over time.

Initial Velocity

The rate of the forward reaction. It is measured quickly before any appreciable amount of P is formed during the Catalytic Step.

Shows that initial velocity is higher as substrate concentration is increased. The relation is not linear but HYPERBOLIC.

Maximum Velocity (Vmax)

Vmax is reached when the increasing of substrate concentration can no longer produce in an increase in the rate of reaction. Each enzyme molecule is working hard to convert S to P.

Michaelis Constant (Km)

Substrate concentration needed to produce one half the maximal velocity of an enzyme catalyzed reaction.

Isozymes

Enzymes that catalyze a given reaction in different tissues but the enzymes have different kinetic parameters.

Often products of different genes.

Glucose enter a cell

A phosphate group is attached to it. It is catalyzed by four isozymes. (hexokinase I, II, III, IV ) The low Km can phosphorylate glucose even when blood concentration is low. The Km is found in the liver, where glucose is stored. Readily responds to glucose. (blood glucose is elevated)

Lineweaver-Burk Plot

Determining reliable values for the kinetic parameters. Reciprocals of velocity and substrate concentration.

If an enzyme is saturated with substrate and is working as fast as possible, adding more enzyme increases the reaction velocity.

However, changing enzyme concentration increases only Vmax and has no influence on Km

A change in Ph can alter these charges (reducing enzyme functions) that may influence directly the active site or some other part of the enzyme that directly affects the active site.

A change in Ph can alter these charges (reducing enzyme functions) that may influence directly the active site or some other part of the enzyme that directly affects the active site. A change in pH may also alter the substrate for an enzyme, which could also influence rate.

Like all chemical reactions, enzymecatalyzed reactions increase in rate if the temperature is increased.

Like all chemical reactions, enzymecatalyzed reactions increase in rate if the temperature is increased. However, since the forces holding the three-dimensional conformation of an enzyme are generally weak, heating too much disrupts the conformation and decreases enzyme activity.

It is also known as the catalytic constant (Kcat), is defined as the maximum number of molecules of substrate converted to product per measure of how fast an enzyme can convert substrate to product.

It is also known as the catalytic constant (Kcat), is defined as the maximum number of molecules of substrate converted to product per measure of how fast an enzyme can convert substrate to product. Since it is considered a measure of the maximum catalytic activity for an enzyme, the enzyme must obviously be saturated with substrate for this to occur.

A better way to express their catalytic efficiency in vivo is to use the expression Kcat/Km , which gives a truer picture of enzyme function under physiological conditions.

A better way to express their catalytic efficiency in vivo is to use the expression Kcat/Km , which gives a truer picture of enzyme function under physiological conditions. The Kcat directly speed with which an enzyme turns substrate into product, whereas the Km is inversely related to the affinity of the enzyme for its substrate.

resemble the normal substrate for an enzyme, for they bind to the active site, but they cannot be changed by the enzyme into product.

resemble the normal substrate for an enzyme, for they bind to the active site, but they cannot be changed by the enzyme into product. They act as mimics, but because of subtle differences between them and the normal substrate they are not chemically altered by the enzyme.

resemble the normal substrate for an enzyme, for they bind to the active site, but they cannot be changed by the enzyme into product. They act as mimics, but because of subtle differences between them and the normal substrate they are not chemically altered by the enzyme. It simply occupies the active site, binding, leaving, binding, and leaving.

Competitive inhibitors compete with the normal substrate for a place on the active site of the enzyme. This inhibition can be overcome by the addition of excess substrate.

Competitive inhibitors compete with the normal substrate for a place on the active site of the enzyme. This inhibition can be overcome by the addition of excess substrate. Competitive inhibitors will not affect the Vmax of the enzyme. However, it will increase the Km because more substrate will be needed to overcome the effects of the competitive inhibitor.

does not resemble the normal enzyme substrate and does not bind to the active site.

does not resemble the normal enzyme substrate and does not bind to the active site. However, when bound to the enzyme, it interferes with its function taking out that enzyme molecule out of commission.

does not resemble the normal enzyme substrate and does not bind to the active site. However, when bound to the enzyme, it interferes with its function taking out that enzyme molecule out of commission. Hence , noncompetitive inhibitors lower Vmax but do not alter Km. Examples are heavy metal ions, knows to cause many health problems because of their tight binding to reactive

Hence , noncompetitive inhibitors lower Vmax but do not alter Km. Examples are heavy metal ions, knows to cause many health problems because of their tight binding to reactive side chains of amino acids in proteins.

Identify first the substrate then the reaction type with the ending ase added. Substrate + reaction(ase).

For example: lactate is reduced through loss of hydrogens and associated electrons (dehydrogenation) fascilitated by specific enzyme. Lactate Dehydrogenase.

Oxidation LEORA (loses electrons oxidation reducing agent) Reduction GEROA (gains electrons reduction oxidizing agent)

Regulation of Enzyme Activity

Each chemical reaction in a cell is catalyzed by a specific enzyme. The rate of an enzyme-catalyzed reaction depends on the concentration of a substrate. Reaction rates can also be controlled by the amount of enzyme protein as well as by the enzymes location within a cell

2 Ways Enzymes could be controlled

Modification by effector molecules Covalent modification

Allosteric Enzymes
Activity is dependent not only in substrates and product concentrations but also on the presence of positive and negative effectors Usually composed of multiple subunits with multiple active sites and are typically placed in metabolic pathways where they can control overall pathway rate.

Allosteric enzymes
These enzymes have sites, other that the active sites wherein effectors can bind. More of a fine tuning type of enzyme activity modulation

Ligands
Molecules that bind to large molecules They may be substrates or products that bind to the active site, as well as allosteric effectors that bind to allosteric sites

Covalent Modification

Can rapidly turn on or off enzyme activity by the covalenty modification of specific amino acid residues in the enzyme protein

Covalent Modification

The most common way to modify the activity of the amino acids is to attatch a phosphate group to the hydroxyl part of the side chains of the amino acids serine, threonine, and tyrosine. This process is called Phosphorylation.

Covalent Modification
Removal of Phosphate group is called Dephosphorylation Protein PhosphorylationDephosphorylation is a common and effective way to alter rapidly and reversibly the activity of key enzymes. Source of the phosphate group is ATP

Covalent Modification
Protein phosphorylation is catalyzed by a class of specific enzymes known as Protein Kinases A class of enzymes known as phosphoprotein phosphatases catalyzes removal of the phosphate groups by hydrolisis. These phosphatases are specific to the amino acid side chain

Purpose: Number of molecules of functional enzyme is proportional to Vmax (enzyme activity). Resulting measurements would be units of reaction velocity : Per unit weight of tissue Per unit amount of protein Per volume of fluid Important in physiological and clinical conditions.

Principles:

Concentration of substrate/s must be high enough to generate a true Vmax. pH of the reaction and the temperature must be standardized. Need to have a simple method for measuring the disappearance of substrate or appearance of product colored substrate or product or can be made to generate a colored complex. International unit (IU)

= Vmax 1 IU = amount of enzyme that converts one

micromole of substrate to product in 1 minute.

BIOCHEMISTRY PRIMER FOR EXERCISE SCIENCE (Houston pp. 17-31)