BABI PANDEY BIOCHEMISTRY VII SEMESTER

Submitted to: Anil Regmi

ENZYMES

OVERVIEW
INTRODUCTION TYPES ISOZYMES

DEFINITION
Enzymes are biocatalysts that are occuring exclusively in

all living cells and catalyze the reactions undergoing within those cells without undergoing any overall change on themselves. Most are proteins Lower the activation energy Increase the rate of reaction Activity lost if denatured May be simple proteins May contain cofactors such as metal ions or organic (vitamins)

PROPERTIES
Higher reaction rate (catalytic power)

Milder reaction conditions

Greater reaction specificity Capacity for regulation

Active site - Specific region in enzyme which interacts with its

substrate. both binding and catalytic reaction occur here. some residues (amino acids) involved in binding substrate others catalyze reaction The shape and the chemical environment inside the active site permits a chemical reaction to proceed more easily

Cofactors
An additional non-protein molecule that is needed by

some enzymes to help the reaction metal ions acts as cofactors--Zn2+,Fe2+,Cu2+,etc Tightly bound cofactors are called Prosthetic groups Cofactors that are bound and released easily are called Coenzymes, coenzymes are organic cofactors Many vitamins are coenzymes

HOLOENZYNE= APOENZYME + COFOCTOR

(INACTIVE PROTEIN) (COENZYME OR METAL ION)

The substrate
The substrate of an enzyme are the reactants that are

activated by the enzyme Enzymes are specific to their substrates The specificity is determined by the active site

ENZYME SUBSTRATE COMPLEX FORMATION

Concept was first conceived by Michaelis and Menten in

1913 Different models have been proposed

Fischers Lock and key Model

Substrate fit into its complementary site on enzyme as key fits

into a lock Ogstons Theory Explains the stereochemical specificity of enzymes There must be at least 3 diff point of interaction
2 for binding & 1 for catalysis

Koshlands Induced Fit Model

A change in the configuration of an enzymes active site (H+ and

ionic bonds are involved). Induced by the substrate.

Lock and Key Model

+
E

+ E +

+
P

ES Complex

Induced Fit Model

P S S P

ES complex
11

E+

ENZYME SPECIFICITY
Group specificity

Absolute specificity
Stereochemical specificity

ACTIVATION ENERGY
Chemical reactions need an initial input of energy = THE

ACTIVATION ENERGY During this part of the reaction the molecules are said to be in a transition state. Increasing the temperature make molecules move faster Biological systems are very sensitive to temperature changes. Enzymes can increase the rate of reactions without increasing the temperature. They do this by lowering the activation energy. They create a new reaction pathway a short cut

E
E Enzyme may be used again P

Enzymesubstrate complex

P
Reaction coordinate

ACTIVATION ENERGY

Factors affecting Enzymes

Substrate concentration

Enzyme concentration
pH Temperature Inhibitors Isoenzymes Cofactors

Substrate concentration
Non-enzymic reactions
Vmax
Reaction velocity Reaction velocity

Enzymic reactions

Substrate concentration

Substrate concentration

Faster reaction but it reaches a

The increase in velocity is

proportional to the substrate concentration

saturation point when all the enzyme molecules are occupied. If you alter the concentration of the enzyme then Vmax will change too.

Enzyme concentration

Reaction velocity

Enzyme concentration

The effect of pH
Optimum pH values

Enzyme activity

Trypsin

Pepsin 1 3 5

7
pH

11

The effect of temperature

Enzyme activity

Q10

Denaturation

10

20 30 40 Temperature / C

50

Inhibitors
Competitive Noncompetitive

CATALYTIC MECHANISM
Acid-Base catalysis

Covalent catalysis
Metal ion catalysis Proximity and orientation effect Preferential binding of the transition state complex Electrostatic catalysis

ENZYME REGULATION
Feedback regulation

Allosteric regulation
Covalent modification Proteolytic activation Enzyme synthesis and breakdown

Allosteric regulation

TYPES

TYPES OF ENZYMES
Metabolic Enzymes They take protein, fat, and carbohydrates and transform them into the proper balance of working cells and tissues. They also remove wornout material from the cells, keeping them clean and healthy. Digestive Enzymes They aid in the digestion of food and the absorption and delivery of nutrients throughout the body. The 3 most important enzymes for digestion are protease, amylase, and lipase. Food Enzymes These enzymes are obtained from the raw vegetables, fruits and other sources of supplements. Similar to the action of digestive enzymes, the food enzymes also help in the digestion of various food nutrients such as the carbohydrates, proteins and fats. The food enzymes are most essential in the maintenance of good health. Bromelain and Papain found in pineapple and papaya respectively and help in the digestion of proteins.

Enzyme Commission (E.C.) Classification

Class EC 1 Oxidoreductases: EC 2 Transferases:
EC 3 Hydrolases:

EC 4 Lyases:

EC 5 Isomerases: EC 6 Ligases:

Reactions catalyzed oxidation/reduction reactions transfer a functional group (e.g.methyl or phosphate group) hydrolysis of various bonds cleave various bonds by means other than hydrolysis and oxidation isomerization changes within a single molecule join two molecules with covalent bonds

ISOZYMES
Isozymes are enzymes that catalyze the same

biochemical reaction but may differ from one another in tissue specificity, developmental regulation or biochemical properties.
Isozymes are encoded by different loci, usually duplicated

genes, as opposed to allozymes, which are different alleles of an enzyme.

Isozymes are important for diagnostic purpose.

Classification of enzyme in blood

Plasma specific Enzymes: functionally active in plasma

eg; blood clotting factors Enzymes of fibrinolysis Secreted enzymes: produced usually in inactive form eg; trypsin, -amylase, chymotrypsin etc. Cellular enzymes: usually synthesized inside the cell--Due to destruction of cell these are excreted to bloodmay be the markersnonspecific marker

Lactate dehydrogenase (LDH)

It can have one of the

following isozymes:
H4 (also called LDH1)

Expressed mostly in the heart + blood cell Expressed mostly in white blood cells Expressed mostly in the lung

H3M (also called LDH2)

H2M2 (also called

LDH3) HM3 (also called LDH4) M4 (also called LDH5)

Expressed mostly in the kidney, placenta Expressed mostly in the liver + muscles

Other isozymes
-amylase P-type S-type Alkaline phosphatase(ALP) Hepatic Bone Placental intestinal

 Acid phosphatase (ACP) Lysosomal Extralysosomal Gama Glutamyl transferase (GGT) Intestine Pancreas Liver Proximal renal tubule

 Lipase Pancreas Lingual glands Gastric, Pulmonary, & Intestinal mucosa SGPT/SGOT

Creatine kinase

CLINICAL SIGNIFICANCE
LDH Increases during MI Increases acute pancreatitis, biliary tract dz, mumps, renal insufficiency, diabetic ketoacidosis

-amylase

Decrease Necrotic pancreatitis, Hepatitis, Severe burns, exocrine pancreatic insufficiency, steatorrhea Aldolase
Increases in skeletal muscle dz,

myopathies

 ALP

Increasea in liver dz, bone dz, growing children Increases in carcinoma of prostate in male,

ACP

pagets dz, hyperparathyroidism, breast cancer

Increases in hepatobiliary dz, intrahepatic or

GGT

Lipase

post hepatic biliary obstruction, liver neoplasm, fatty liver, infectious hepatitis, alcoholic hepatitis, pancreatitis, pancreatic malignancy Acute pancreatitis, obstruction of pancreatic duct by calculi, carcinoma of pancreas, renal dz, reduced GFR in patient

 SGPT

Increases in hepatocellular injury, it often

SGOT

suggest the existence of other medical problems such as viral hepatitis, diabetes, congestive heart failure, liver damage, bile duct problems, infectious mononucleosis, or myopathy. ALT is a more specific indicator of liver inflammation than AST, as AST may be elevated also in diseases affecting other organs, such as myocardial infarction, acute pancreatitis, acute hemolytic anemia, severe burns, acute renal disease, musculoskeletal diseases, and trauma.[18]

 creatine kinase

creatine kinase is assayed

in blood tests as a marker of myocardial infarction (heart attack), rhabdomyolysis (severe muscle breakdown), muscular dystrophy, the autoimmune myositides and in acute renal failure.

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