ENZYMES

ENZYMES
• An enzyme is a protein that acts as a catalyst for a biological
reaction.
• Most enzymes are specific for substrates while enzymes
involved in digestion such as papain attack many substrates
 ENZYMES
• Enzymes are complex organic catalysts of highly specific action that are of vital
importance in biological processes.
• They are manufactured by living cells,but are able to function without the
presence of these cells.
• They modify the speed of a reaction without being used up or appearing as one
of the end products of the reaction.
 NATURE OF ENZYMES:

• Enzymes are proteins of low molecular weight.

• They are coagulated and so become inactive by high temperature, alcohol, salts, heavy
metals,and alkaloidal reagents.
• In general,they are soluble in water.
• Most reactions of living matter are accelerated by enzymes.
• Enzymes are responsible for the reactions like oxidation, reduction, hydrolysis, synthesis
and many others which are necessary during digestion, metabolism, respiration, energy
transfers and release.
ENZYMES ARE THE AGENTS
OF METABOLIC FUNCTION
regulatory enzymes capable of
sensing the momentary metabolic
needs of the cell and adjusting
their catalytic rates
accordingly.
 COFACTORS

• In addition to the protein part, many enzymes also have a nonprotein part called a
cofactor
• The protein part in such an enzyme is called an apoenzyme,and the combination
of apoenzyme plus cofactor is called a holoenzyme. Only holoenzymes have
biological activity; neither cofactor nor apoenzyme can catalyze reactions by
themselves
• A cofactor can be either an inorganic ion or an organic molecule, called a
coenzyme
• Many coenzymes are derived from vitamins,organic molecules that are dietary
requirements for metabolism and/or growth
 SOME C O M M O N ENZYMES A N D THEIR ACTIVITIES
ENZYMES WHERE FOUND SUBSTRATE END PRODUCTS
I. Hydrolytic Enzymes:
CARBOHYDRASES
Ptyalin or salivary amylase Saliva Starch & glycogen Maltose
Amylopsin or Pancreatic Pancreatic juice Starch & glycogen Maltose

Maltase Intestinal juice Maltose Glucose

Lactase Intestinal juice Lactose Glucose & galactose

Sucrase Intestinal juice Sucrose Glucose & fructose

ESTERASES OR LIPASES
Gastric Lipase Gastric juice in Emulsified fats Fatty acid & glycerol
Stomach
Streapsin or Pancreatic lipase Pancreatic juice Fats Fatty acid & glycerol

Phosphatases Tissues Hexophosphate Hexose & H3PO4

ENZYMES WHERE FOUND SUBSTRATE END PRODUCTS

PROTEASES:
. Pepsin Gastric juice Proteins Proteases &
Peptoses
Trypsin or Creatic Pancreatic juice Proteins, Polypeptides
Proteinase proteoses,
peptones
Pancreatic juice Proteins, Polypeptides
Chymotrysin proteoses,
peptones
Amino polypeptidases Intestinal juice Polypeptides Peptides &
& Carboxypeptidases amino acids

Dipeptidases Intestinal & Dipeptides Amino acids

Pancreatic juice
II. OXIDO-REDUCTASES:
Dehydrogenases Animal tissues Hydrogen donors Oxidized hydrogen
acceptor

Oxidases Plants & animal Carbohydrate proteins, fats in CO2 & H2O
Tissues tissues

Peroxidases Plant & animal Organic peroxides Oxygen

Tissues
. Catalases Plant & animal Hydrogen peroxides Water & oxygen
Tissues
III. FERMENTING ENZYMES:
. Zymase Yeast Monosaccharides CO2 and ethyl alcohol

Lactic acidase Lactic acid Lactose Lactic acid

Bacteria
IV. COAGULATING ENZYMES:
Rennin/Rennet Gastric juice Milk or casein Paracasein
 D EFINITION OF TERMS
• Zymogen or proenzyme- is the inactive form of the enzymes. Ex. pepsinogen is the
inactive form of pepsin; trypsinogen is the inactive form of trypsin.
• Activator or kinase- is a substance which activates the proenzyme. If the activator is an
organic substance it is called a kinase. Ex. HCl in the gastric juice activates pepsinogen
into pepsin, trypsinogen is activated by enterokinase in the intestinal juice into trypsin.
• Antienzymes- substances which inhibit enzyme activity. They are found in the lining of
the stomach and intestines, inhibit the action of pepsin on the stomach protein.
• Coenzymes- substance or electrolytes, or non-electrolytes which are required by a
particular enzyme for its activity. Ex. Bile salts, chloride ions.
 HO W D O ENZYMES W ORK?
EXAMPLE: CITRATE SYNTHASE
• Citrate synthase catalyzes a mixed Claisen condensation of acetyl CoA and
oxaloacetate to give citrate
The Claisen condensation is a carbon–carbon bond forming reaction that
occurs between two esters or one ester and another carbonyl compound in
the presence of a strong base, resulting in a β-keto ester or a β-diketone.
• Normally Claisen condensation requires a strong base in an alcohol solvent
but citrate synthase operates in neutral solution
THE STRUCTURE OF CITRATE SYNTHASE
• D etermined by X -ray crystallography
• Enzyme is very large compared to substrates, creating a
complete environment for the reaction
 PROTEIN DENATURATION
• The tertiary structure of a globular protein is the result of
many intramolecular attractions that can be disrupted by a
change of the environment, causing the protein to become
denatured
• Solubility is drastically decreased as in heating egg white,
where the albumins unfold and coagulate
• Enzymes also lose all catalytic activity when denatured
 U N D E R S TA N D I N G CATALYSIS

H O W D O ENZYMES
W O R K A S C ATA LYSTS?
Lock-and-key model

portrays an enzyme as conformationally rigid

and able to bond only to substrates that exactly
fit the active site.
The INDUCED-FIT MODEL states a substrate binds to an active site and both
change shape slightly, creating an ideal fit for catalysis. Enzymes promote
chemical reactions by bringing substrates together in an optimal orientation, thus
creating an ideal chemical environment for the reaction to occur.
Substance which promotes
the activity of catalyst is
called “promotor” or
“activator”. it is also called
“catalys for a catalyst”.
t such phenomenon is
catalyst activation.
Examples: Hydrogenation
of vegetable oil to
vegetable ghee is
accelerated by nickel.
 CLINICAL SIGNIFICANCE

• Today many competitive inhibitors are used as drugs.

• Sulfanilamide, a sulfur drug, is an antibacterial agent
because it is a competitive inhibitor of an enzyme-
catalyzed reaction using p-aminobenzoic acid in the
synthesis of folic acid.
• Folic acid is a water-soluble vitamin.
Sulfanilamide is apparently not harmful to
man because folic acid is not synthesized
by man but obtained in the diet.
• Apparently bacteria can’t absorb sufficient
folic acid from the host and are
susceptible to inhibition by such drugs.
• Methotrexate (amethopterin) is a competitive
inhibitor of dihydrofolate reductase because its
structure is similar to dihydrofolate
• Methotrexate is used in cancer chemotherapy
because it blocks tetrahydrofolate synthesis
which is essential to the biosynthesis of the
thymine mononucleotide. Rapidly dividing
cancer cells which are synthesizing D N A are
more susceptible to methotrexate than slower
growing normal cells.
 ALTERNATIVE MEDICINE USING ENZYMES
Rating Health Concerns
Low back pain (chymotrypsin,trypsin)
Pancreatic insufficiency (including pancreatitis)
Sprains and strains (chymotrypsin, trypsin)
Indigestion (Lipase )
O steoarthritis (bromelain,trypsin,rutosid combination)
Tendinitis (proteolytic enzymes)
Acne Rosacea
Chronic candidiasis
Food allergies
Gastroesophageal reflux disease (GERD)
Low back pain (papain)
Sprains and strains (papain)
SOURCE: http://www.peacehealth.org
IN INDUSTRIES

• Cellulase is the major industrial enzyme

used to soften and fade jeans
(environment-friendly and effective)
• http://www.europabio.org/documents/JEANS.p
df
 DAIRY PRODUCTS INDUSTRIES
• Source:www.biotech.about.com.
• Biotechnology gives many industrial applications that
result in biotech products that we use everyday at
home
• Some of these are food science applications that
utilize enzymes to produce or make improvements in
the quality of different foods.
• In the dairy industry, some enzymes are required for
the production of cheeses, yogurt and other dairy
products, while others are used in a more specialized
fashion to improve texture or flavour
 CH EESE-MAKING

• Cheese consists of proteins and fat

from milk,usually the milk of cows,
buffalo,goats, or sheeps.It is
produced by coagulation of the milk
protein casein.Typically,the milk is
acidified and addition of the enzyme
rennet causes coagulation.
•
COENZYMES
 C O E N ZYMES
• There are other groups that contribute to the reactivity of enzymes
beside amino acid residues.
• These groups are called cofactors - chemicals required by
apoenzymes (inactive) to become holoenzymes (active).
• There are two types of cofactors:
1) essential ions - metal ions -inorganic
2)coenzymes - organic molecules that act as group-transfer reagents
(accept or donate groups)- can also be H + and/or e-
Both provide reactive groups not found on a.a.side chains.
1) METABOLITE COENZYMES

• synthesized by common metabolites

• include nucleoside triphosphates
• most abundant isATP, but also include uridine diphosphate glucose (UDP-
glucose) and S-adenosylmethionine
• ATP can donate all of its three phosphoryl groups in group-transfer reactions
• S-adenosylmethionine (SAM) can donate its methyl group in biosynthetic
reactions.
• UDP-glucose is a source of glucose for synthesis of glycogen in animals and
starch in plants.
2) vitamin-derived coenzymes
• Vitamins are required for coenzyme synthesis and must be supplied in the
diet
• Lack of particular vitamins causes disease
• There are two catagories of vitamins:
1)water-soluble - B vitamins and vit.C- required daily in diet, excess
excreted in urine
2) lipid-soluble - vitaminsA, D,E,K
Intake must be limited Stored
in fat
 B VITAMINS A N D THEIR C O E N Z Y M E S VITAMINB3 N I A C I N

nicotinic acid → nicotinamide

• Get niacin in enriched cereals,meat,legumes.
• NAD + and NADP + are the coenzymes (cosubstrates).
• N A D + consists of 2 5’ribonucleotides (AMP and nicotinamide monomucleotide) joined by a
phosphoanhydride linkage.

• For NADP + ,have a phosphoryl group on 2’-oxygen.

• Both coenzymes act as cosubstrates for dehydrogenases → catalyze the oxidation of substrates by transfer
of 2e- and 1H+ →N A D H and N A DPH.
 VITAMIN B 2 (RIBOFLAVIN)

• Coenzymes are flavin mononucleotide (FMN) and flavin adenine dinucleotide (FAD).
• Riboflavin found in milk, whole grains, liver.
• The coenzymes serve as prosthetic groups involved in 1e- or 2e- transfers.
FAD + 2e- + 2H+ → FADH 2
FMN + 2e- + 2H+ → FMNH 2
• Enzymes that require FAD or FMN are called flavoenzymes or flavoproteins.
• Can actually donate 1 or 2 e- at a time → form partially oxidized compound when only 1e- is donated → relatively
stable.
 V IT AM IN B 1 (THIAMINE)

• Structure: pyrimidine ring and positively charged thiazolium ring.

• Found in husks of rice and other cereals,liver,meat,particularly
pork.
• Deficiency in thiamine causes beriberi - extensive nervous system
and circulatory system damage,muscle wasting,edema.
• Coenzyme form is thiamine pyrophosphate (TPP) - synthesized by
transfer of pyrophosphoryl group from ATP via thiamine
pyrophosphate synthetase.
• Used primarily in decarboxylases as a coenzyme.
 V IT AM IN B6 FAMILY
• pyridoxine, pyridoxal, pyridoxamine are the vitamins.
• Act as prosthetic groups.
• Formed by the following reaction:
pyridoxine + ATP → pyridoxine 5’phosphate → pyrodoxal 5’ phosphate (PLP).
• Lack of B6 results in defects in protein metabolism.
 B IO T I N

• Synthesized by intestinal bacteria.

• Prosthetic coenzyme is called biocytin - covalently linked to Lys residue in
active site.
• Involved in carboxyl group transfer reactions and ATP-dependent
carboxylations.

• E.g.pyruvate carboxylase
pyruvate + HCO 3 - →
oxaloacetate

• Binds to HC O 3- and acts as a C O 2 carrier

FOLI C A C I D O R FOLATE

• Found in green leafy vegetables,liver,yeast.

• Coenzyme form is tetrahydrofolate.
• Used by enzymes that transfer 1-C units as methyl groups (CH 3-).
• Another folate coenzyme is tetrahydrobiopterin - used in hydroxylases
 VITAMIN B5: P A N T H O T H E N I C A C I D

• Used in coenzymeA formation.

• Reactive center is -SH group
• Key in all acyl-group transfers
• Coenzyme form is phosphopantethine - added to serine
residue of protein --> acyl carrier protein (ACP) -->
important in fatty acid synthesis.
 VITAMIN B12 O R C O B A L A M I N

• Found in organ meat (kidney and liver).

• It is a prosthetic coenzyme.

• Ring structure similar to heme, with cobalt atom in center.

• Involved in molecular rearrangements.
• Deficiency in B12 results in pernicious anemia (decreased production of blood cells from
bone marrow).
VITA M I N C O R A S C O R B I C A C I D

• Found in fresh fruit and vegetables.

• Participates in hydroxylation reactions, e.g.collagen synthesis.
• Deficiency causes scurvy.
 LIPID V ITA M I N S /VITA M I N A O R RETIN O L

• Is a 20 carbon lipid molecule.

• Found in carrots, yellow vegetables, liver, egg yolk, milk products.
carotene → vitaminA
Exists in three forms:
1) retinol and 2) retinoic acid - bind to intracellular protein receptors → regulates
gene expression
2) retinal - prosthetic group of rhodopsin
 VITAMIN D

• Exists as several lipids;

• 1) D3 - made in skin exposed to sunlight.
• 2) D2 - additive in fortified milk
• Deficiency causes rickets in children or osteomalacia in
adults --> insufficient Ca phosphate deposition in bone.
VITA M I N E O R A -TO C O P H E RO L

• an antioxidant that scavenges free radicals.

 VITAMIN K O R P H Y L L O Q U I N O N E

• Found in plants.
• Required for synthesis of
proteins involved in
blood coagulation.
 UBIQUINONE OR COENZYME Q

• Ring with hydrophobic tail --> inserted into membranes.

• Transports e- between enzyme complexes in inner
mitochondrial membrane.
• Related molecule is plastiquinone - found in thylacoid
membrane of chloroplasts.
CYTOCHROMES

• Hemo-containing protein coenzyme Fe3+ <--> Fe2+.

• Classified as a,b,c based on absorption spectra.
• Transfers e-.