FACTORS AFFECTING ENZYME ACTIVITY

Velocity or rate of enzymatic reaction is assessed by the rate of change in concentration of substrate or product at a given time duration. Various factors which affect the activity of enzymes include:

1. Substrate concentration

2. Enzyme concentration

3. Product concentration

4. Temperature 5. Hydrogen ion concentration (pH)

6. Presence of activators

7. Presence of inhibitor

Effect of substrate Concentration :  Reaction velocity of an enzymatic process increases with constant enzyme concentration and increase in substrate concentration.

Effect of enzyme Concentration: As there is optimal substrate concentration, rate of an enzymatic reaction or velocity (V) is directly proportional to the enzyme concentration.

Effect of product concentration In case of a reversible reaction catalyzed by a enzyme, as per the law of mass action the rate of reaction is slowed down with equilibrium. So, rate of reaction is slowed, stopped or even reversed with increase in product concentration

Effect of temperature: Velocity of enzymatic reaction increases with temperature of the medium which they are most efficient and the same is termed as optimum temperature.

Effect of pH: Many enzymes are most efficient in the region of pH 6-7, which is the pH of the cell. Outside this range, enzyme activity drops off very rapidly. Reduction in efficiency caused by changes in the pH is due to changes in the degree of ionization of the substrate and enzyme.

Highly acidic or alkaline conditions bring about a denaturation and subsequent loss of enzymatic activity

Exceptions such as pepsin (with optimum pH 1-2), alkaline phosphatase (with optimum pH 9-10) and acid phosphatase (with optimum pH 4-5)

Presence of activators Presence of certain inorganic ions increases the activity of enzymes. The best examples are chloride ions activated salivary amylase and calcium activated lipases.

Effect of Inhibitors The catalytic enzymatic reaction may be inhibited by substances which prevent the formation of a normal enzyme-substrate complex. The level of inhibition then depends entirely upon the relative concentrations of the true substrate and the inhibitor

PHOSPHOLIPIDS

These are complex or compound lipids containing phosphoric acid, in addition to fatty acids, nitrogenous base and alcohol 

There are two  classes of phospholipids

1. Glycerophospholipids (or phosphoglycerides) that contain glycerol as the alcohol.

2. Sphingophospholipids (or sphingomyelins) that contain sphingosine as the alcohol

Glycerophospholipids

Glycerophospholipids are the major lipids that occur in biological membranes. They consist of glycerol 3-phosphate esterified at its C1 and C2 with fatty acids. Usually, C1 contains a saturated fatty acid while C2 contains an unsaturated fatty acid.

In glycerophospholipids, we refer to the glycerol residue (highlighted red above) as the "glycerol backbone."

Glycerophospholipids are Amphipathic

Glycerophospholipids are sub classified as

1. Phosphatidylethanolamine or cephalin also abbreviated as PE is found in biological membranes and composed of ethanolamine bonded to phosphate group on diglyceride.

2. Phosphatidylcholine or lecithin or PC which has chloline bonded with phosphate group and glycerophosphoric acid with different fatty acids like palmitic or hexadecanoic acid, margaric acid, oleic acid. It is a major component of cell membrane and mainly present in egg yolk and soy beans.

3. Phosphatidic acid (phosphatidate) (PA)

It consists of a glycerol with one saturated fatty acid bonded to carbon-1 of glycerol and an unsaturated fatty acid bonded to carbon-2 with a phosphate group bonded to carbon-3.

4.Phosphatidylserine (PS)

This phospholipid contains serine as an organic compound with other main components of phospholipids. Generally it found on the cytosolic side of cell membranes.

5. Phosphoinositides

It is a group of phospholipids which are negatively charged and act as a a minor component in the cytosolic side of eukaryotic cell membranes. On the basis of different number of phosphate groups they can be different types like phosphatidylinositol phosphate (PIP), phosphatidylinositol bisphosphate(PIP2) and phosphatidylinositol trisphosphate (PIP3). PIP, PIP2 and PIP3 and collectively termed as phosphoinositide.

6. Cardiolipin :

lt is so named as it was first isolated from heart muscle. Structurally, a cardiolipin consists of two molecules of phosphatidic acid held by an additional glycerol through phosphate groups. lt is an important component of inner mitochondrial membrane. Cardiolipin is the only phosphoglyceride that possesses antigenic properties.

PROPERTIES OF TRIACYLGTYCEROLS

1. Hydrolysis : Triacylglycerols undergo stepwise enzymatic hydrolysis to finally liberate free fatty acids and glycerol.

The process of hydrolysis, catalysed by lipases is important for digestion of fat in the gastrointestinal tract and fat mobilization from the adipose tissues.

2. Saponification : The hydrolysis of triacylglycerols by alkali to produce glycerol and soaps is known as saponification.

3.Rancidity: Rancidity is the term used to represent the deterioration of fats and oils resulting in an unpleasant taste. Fats containing unsaturated fatty acids are more susceptible to rancidity.

Hydrolytic rancidity occurs due to partial hydrolysis of triacylglycerols by bacterial enzymes.

Oxidative rancidity is due to oxidation of unsaturated fatty acids.

This results in the formation of unpleasant products such as dicarboxylic acids, aldehydes, ketones etc.

Antioxidants : The substances which can prevent the occurrence of oxidative rancidity are known as antioxidants.

Trace amounts of antioxidants such as tocopherols  (vitamin E), hydroquinone, gallic acid and c,-naphthol are added to the commercial preparations of fats and oils to prevent rancidity. Propylgallate, butylatedhydroxyanisole (BHA)  and butylated hydroxytoluene (BHT) are the antioxidants used in food preservation.

Lipid peroxidation in vivo: In the living cells, lipids undergo oxidation to produce peroxides and free radicals which can damage the tissue. .

The free radicals are believed to cause inflammatory diseases, ageing, cancer , atherosclerosis etc

Iodine number : lt is defined as the grams (number)  of iodine absorbed by 100 g of fat or oil. lodine number is useful to know the relative

unsaturation of fats, and is directly proportional to the content of unsaturated fatty acids

Determination of iodine number will help to know the degree of adulteration of a given oil

Saponification number : lt is defined as the mg  (number) of KOH required to hydrolyse (saponify) one gram of fat or oiL

Reichert-Meissl (RM)  number: lt is defined as the number of ml 0.1 N KOH required to completely neutralize the soluble volatile fatty acids distilled from 5 g fat. RM number is useful in testing the purity of butter since it contains a good concentration of volatile fatty acids (butyric acid, caproic acid and caprylic acid).

Acid number : lt is defined as the number of mg of KOH required to completely neutralize free fatty acids present in one gram fat or oil. In normal circumstances, refined oils should be free from any free fatty acids.

Sphingosine is an amino alcohol present in sphingomyelins (sphingophospholipids).  They do not contain glycerol at all.

Sphingosine is attached by an amide linkage to a fatty acid to produce ceramide. The alcohol group of sphingosine is bound to phosphorylcholine in sphingomyelin structure. .

Sphingomyelins are important constituents of myelin and are found in good quantity in brain and nervous tissues.

Glycogen Metabolism

The formation of glycogen from glucose is called Glycogenesis

Glycogen is a polymer of glucose residues linked mainly by a(1→ 4)  glycosidic linkages. There are a(1→6) linkages at branch points. The chains and branches are longer than shown. Glucose is stored as glycogen predominantly in liver and muscle cells

Glycogen Synthesis

Uridine diphosphate glucose (UDP-glucose) is the immediate precursor for glycogen synthesis. As glucose residues are added to glycogen, UDP-glucose is the substrate and UDP is released as a reaction product. Nucleotide diphosphate sugars are precursors also for synthesis of other complex carbohydrates, including oligosaccharide chains of glycoproteins, etc.

UDP-glucose is formed from glucose-1-phosphate and uridine triphosphate (UTP)

glucose-1-phosphate + UTP → UDP-glucose + 2 P_i

Cleavage of PPi is the only energy cost for glycogen synthesis (1P bond per glucose residue)

Glycogenin initiates glycogen synthesis. Glycogenin is an enzyme that catalyzes glycosylation of one of its own tyrosine residues.

Physiological regulation of glycogen metabolism

Both synthesis and breakdown of glycogen are spontaneous. If glycogen synthesis and phosphorolysis were active simultaneously in a cell, there would be a futile cycle with cleavage of 1 P bond per cycle

To prevent such a futile cycle, Glycogen Synthase and Glycogen Phosphorylase are reciprocally regulated, both by allosteric effectors and by covalent modification (phosphorylation)

Glycogen catabolism (breakdown)

Glycogen Phosphorylase catalyzes phosphorolytic cleavage of the →(1→4) glycosidic linkages of glycogen, releasing glucose-1-phosphate as the reaction product.

Glycogen (n residues) + P_i → glycogen (n-1 residues) + glucose-1-phosphate

The Major product of glycogen breakdown is glucose -1-phosphate

Fate of glucose-1-phosphate in relation to other pathways:

Phosphoglucomutase catalyzes the reversible reaction:

Glucose-1-phosphate → Glucose-6-phosphate

TRIGLYCEROL

Triacylglycerols (formerly triglycerides) are the esters of glycerol with fatty acids. The fats and oils that are widely distributed in both  plants and animals are chemically triacylglycerols.

They are insoluble in water and non-polar in character and commonly known as neutral fats.

Triacylglycerols are the most abundant dietary lipids. They are the form in which we store reduced carbon for energy. Each triacylglycerol has a glycerol backbone to which are esterified 3 fatty acids. Most triacylglycerols are "mixed." The three fatty acids differ in chain length and number of double bonds

Structures of acylglycerols :

Monoacylglycerols,  diacylglycerols and triacylglycerols, respectively consisting of one, two and three molecules of fatty acids esterified to

a molecule of glycerol

Lipases hydrolyze triacylglycerols, releasing one fatty acid at a time, producing  diacylglycerols, and eventually glycerol

Glycerol arising from hydrolysis of triacylglycerols is converted to the Glycolysis intermediate dihydroxyacetone phosphate, by reactions catalyzed by:
(1) Glycerol Kinase
(2) Glycerol Phosphate Dehydrogenase

Free fatty acids, which in solution have detergent properties, are transported in the blood bound to albumin, a serum protein produced by the liver.
Several proteins have been identified that facilitate transport of long chain fatty acids into cells, including the plasma membrane protein CD36