ENZYMES ACTIVATION AND DEACTIVATIONNovember 19th, 2012

Enzymes are Not Consumed in Reaction

https://www.youtube.com/watch?v=0XjyAkeQJag&feature=related

Factors Effecting Enzymes Enzymes are not perfect They respond to environmental

conditions They work depending on various

factors which are? ________, ___________, _____________ Changing these factors

effects? _____________

Enzyme under Stress

pH An decrease in pH, increases the [H+]

ion concentration in solution An increase in pH, increases the [OH-]

ion concentration in solution These ions interfere with hydrogen

bonds and ionic bonds Changing the conformation of the

enzymes specifically the active site Activity of enzyme is affected

Optimal pH Different enzymes have different optimum pHs At optimum pH the active site is the shape

most complementary to the shape of their Substrate

At optimum pH, the rate of reaction is highest Large changes in pH can cause

enzymes to Denature and permanently loose their function

Temperature Effects Enzymes Generally, enzymes have a narrow

range of temperature they work in, Why?

At the optimal temperature enzymes are most active

Increase Temperature

What increases? energy Makes the substrate more active in

solution. So? More chances of substrate colliding with

active site. Makes the enzyme more flexible. Puts strain on weaker bonds. Pass a certain point enzymes denature.

What does it mean to denature? active site changes

Decrease Temperature Makes the enzyme less flexible, Pass a certain point enzymes do not

function properly Not enough energy present

Taq Polymerase Enzyme comes from Thermus Aquaticus,

a species that thrive in hot springs and heat vents.

Functions at high temperature Used in Polymerase Chain Reaction (PCR) Can make multiple copies of a DNA sample

using only a small amount. PCR can be used for a forensic investigation,

genetic diseases, drug discovery and detection of pathogens

Temperature Curve Various thermophillic organisms have

their own type of DNA polymerase such as Pfu Polymerase (Pyrococcus furiosus) versus Taq Polymerase

Regulation of Temperature Endotherms can maintain body temperature Heat is produced and

regulated by the body, How? Ectotherms do not maintain a body

temperature Less sensitive to changes in body temperature Endothermic organisms are mammals, birds

and some fish Most enzymes in the human body have an

optimal temperature of 37°C

Concentration What is concentration?

Will the concentration of substrate the rate of reaction?

Will the concentration of enzyme effect the rate of reaction?

Substrate Concentration Adding more substrate increases rate of

the reaction More substrate molecule collide with

active site At a certain point adding more substrate

has no more effect. Enzyme active site is saturated

Enzyme Concentration If an enzyme is saturated what can you

do? Increasing enzyme concentration,

increases the rate of reaction Why does the graph level off?

ENZYME REGULATIONNovember 20th & 21st, 2012

Biochemical Process

http://www.iubmb-nicholson.org/animaps.html

Enzyme Regulation There are enzymes for

each specific reaction of the human body

There is a need to control enzyme activity

Regulation is efficiency

Enzymes can be activated and inhibited

Road Map

Aspirin Cyclooxygenase 2 (COX2) makes prostaglandins These chemical are involved in inflammation Inflammation is felt as pain and swelling in body Aspirin reacts with the amino acid serine irreversibly,

blocking the active site, substrate can not bind Other pain killers such as ibuprofen (Advil) bind less

strongly, are reversible bound

Inhibition Enzyme inhibitors are substances that

interfere with catalysis Inhibitors slow down the rate of reaction Inhibitors can be reversible or irreversible Irreversible inhibition – halts enzymatic reaction

permanently Reversible inhibition – slows down the reaction

temporarily Inhibitors can act in a competitive or non

competitive form and interfere with the reaction

Competitive Inhibition Competitive inhibition: Enzyme

inhibitors prevent the formation of Enzyme-Substrate complexes because they have a similar shape to the substrate molecule.

Prevents enzyme from carrying out reaction it is suited for

Enzyme with Active Site Specific for Substrate

Substrate

Inhibitor

Inhibitor Competes with Substrate for the Active Site

Competitive Inhibition Inhibitor has a different shape than the substrate

but complements the active site Inhibitor does not react since it has different

structure than the substrate. Reaction rate is decreased since fewer substrate

molecules can bind to the enzyme Inhibition is typically temporary, the inhibitor

eventually leaves the active site Inhibition depends on the relative

concentrations of substrate and inhibitor, both compete for place in enzyme active site

Methanol Poisoning Methanol if ingested is oxidized to

formaldehyde and formic acid Attack on the optic nerve causes blindness. Methanol found in engine fuel,

solvents, window cleaner, andantifreeze

Source: http://curriculum.toxicology.wikispaces.net/2.2.5.2.5+Methanol

Ethanol Competes with Methanol

Ethanol competitively inhibits the oxidation of methanol by Alcohol Dehydrogenase

Ethanol is oxidized in preference to methanol

Oxidation of methanol is slowed down

Toxic by-products do not have chance to accumulate.

Source: http://curriculum.toxicology.wikispaces.net/2.2.5.2.5+Methanol

Pennicillin Pennicillin, an antibiotic, works against disease causing

bacteria Stops cell wall cross-linking permanently Inactivates transpeptidase, used to build cross-linked

peptidoglycan layer in the membrane The cross-linking peptide chains have repeats of D-Alanine Pennicillin also has a repeat of D-Alanine-D-Alanine

E.Coli cells can not grow and die

Succinate Dehydrogenase Inhibitor Succinate Dehydrogenase catalyzes the conversion

of succinate to fumerate, an important biochemical reaction in cellular respiration.

Malonate inhibits this reaction competitively Used to find

active sitechemistry

Used to study inborn errors of metabolism

Non-Competitive Inhibition Non-competitive inhibition: enzyme

inhibitors prevent the formation of Enzyme-Product Complexes.

Inhibitors prevent the substrate to react and form into product

Non-competitive inhibitors bind to a site other than the Active Site

Binding causes conformational changes that change the tertiary structure of the enzyme

Thus, enzyme can not catalyze reaction

Non-Competitive Inhibition

Substrate

Non-competitive Inhibitor

Enzyme Active Site Complementary to Substrate

Non-Competitive Reaction Since they do not compete with

substrate molecules, non-competitive inhibitors are not affected by substrate concentration.

Many non-competitive inhibitors are irreversible and permanent, and effectively denature the enzymes which they inhibit.

However, there are a lot of non-permanent and reversible non-competitive inhibitors that are vital in controlling metabolic functions in organisms.

Cyanide Poisoning Another enzyme found in cellular respiration is

cytochrome oxidase, one of the most important enzymes in the electron transport chain of reactions that occurs in the mitochondria inner membrane

Here oxygen is reduced and 34 ATP molecules are made.

Cyanide Poisoning Cyanide acts as a non-competitive inhibitor for

cytochrome oxidase complex Cyanide does not compete for the active sites of the

enzyme because it has no similarity to the substrate cytochrome

Cyanide attaches to another site on the enzyme and disrupts the enzyme's shape.

This brings the electron transport chain to a halt No energy can be derived out of respiration Hydrogen cyanide inhibits metal-containing enzymes in

the body, such as cytochrome c-oxidase, which contains iron

Irons in Enzymes Chemical catalyst are usually metals Many enzymes get their ability to

catalyze reactions due to metals found in the active site

One common metal used is iron Fe2+ that is found in a protoporphyrin ring

Ferrochelatase

Ferrochetalase inserts iron into protoporphyrin rings

Lead forms covalent bonds with the sulphydryl side chains of the amino acid cysteine in the enzyme and prevents catalytic activity

The binding of the heavy metal shows non-competitive inhibition because the substrate still has access.

Chymotrypsin Chymotrypsin is an enzyme which

hydrolyzes peptides bonds In its active site there are three amino

acids Histidine57, Serine195 and Asparagine102 known.

Hydrogen Ion Inhibits Chymotrypsin These amino acids allow for the substrate

to be cleaved. By lowering pH, amino acids in the active

site no longer accept hydrogen proton since Asp102 becomes protonated (hydrogens added)

Hydrogen ion acts as a non-competitive inhibitor by preventing catalysis but do not prevent the substrate from binding to the active site.

Biochemical Pathway

A biochemical pathway is a series of step reactions leading to a product

Enzymes lie in biochemical pathways There are specific enzymes for each

reaction step Metabolism is a sum of biochemical

pathways and is made of anabolic and catabolic processes

Need to Regulate

There are so many pathways that are incorporated in the metabolic system of the human body

An efficient process is needed to regulate the use of resources and ensure that only what is required is being produced or broken down

Enzymes can be regulated by the ability to be activated and deactivated when needed

Allosteric Enzyme Regulation

An allosteric site, a site away from the active site,can bind molecules to change conformation of the enzyme.

At the allosteric site for an inhibitor, binding of an inhibitor causes a conformational change such that the active sites of an enzyme are non complementary to the substrate.

An activator can bind to its allosteric site to open or improve the fit between substrate and enzyme.

Feedback Inhibition In a biochemical pathway, by controlling

an earlier step, the next series of reaction steps can be controlled

Usually the end product in a chain of reactions is an inhibitor of an earlier enzyme in the chain to stop the creation of more product

Process is self-regulating and cell resources are not wasted by making more product than needed

Feedback Inhibition Using an Allosteric Site

Regulation of Glycolysis Glycolysis is biochemical process where

glucose is broken down to pyruvate Pyruvate is used in mitochondria in the

process of aerobic respiration to derive ATP Pyruvate kinase is the enzyme that converts

phosphenolpyruvate to pyruvate in glycolysis This enzyme is the third regulated enzyme of

glycolysis ATP and alanine act as allosteric inhibitors of

pyruvate kinase

Feedback Inhibition of Pyruvate