Enzymes are biological catalysts that facilitate chemical processes in living organisms. They are primarily proteins, although some RNA molecules can also function as enzymes (known as ribozymes). It decreases the activation energy required for a reaction, allowing it to proceed more quickly and efficiently.
Table of Contents
Classification of Enzymes
Oxidoreductases: Protein catalysts that facilitate the transfer of electrons between molecules, often involved in oxidation-reduction reactions.
Example: Cytochrome oxidase.
Transferases: Protein catalysts that transfer functional groups (such as methyl or phosphate) from one molecule to another.
Example: Alanine deaminase.
Hydrolases: Protein catalysts that catalyze the breakdown of molecules by adding water (hydrolysis).
Example: Lipase.
Lyases: Protein catalysts that add or remove groups to/from double bonds, resulting in the formation or breaking of double bonds.
Example: Oxalate decarboxylase.
Isomerases: Protein catalysts that rearrange the atoms within a molecule to form isomers.
Example: Glucose-phosphate isomerase.
Ligases: Protein catalysts that join two molecules together, often using energy from ATP.
Example: DNA ligase.
Sources of Enzymes
Enzymes are present in all living organisms. They can be produced naturally or through special technique that produces specialized protein catalysts.
Naturally occurring enzymes
Plants: Protein catalysts such as amylase (found in saliva and helps break down starch), bromelain (found in pineapple and helps break down protein), and papain (found in papaya and helps break down protein).
Animals: Protein catalysts like pepsin (found in the stomach and helps break down protein), trypsin (found in the pancreas and helps break down protein), and lipase (found in the pancreas and helps break down fats).
Microorganisms: Protein catalysts like lactase (found in some bacteria and helps break down lactose), and penicillin acylase (found in some fungi and helps produce penicillin).
Enzymes produced using special techniques
Genetic engineering: Scientists can change organisms’ genes to create certain protein catalysts. This is how many industrial protein catalystsare produced.
Chemical synthesis: Some protein catalystscan be created in the lab using chemicals. This is a more complicated method than genetic engineering, but it can manufacture protein catalysts that do not occur naturally.
Mechanism of Enzyme Action
Enzymes are biological catalysts that accelerate chemical reactions by lowering the activation energy needed for them to occur. They accomplish this by attaching to substrates (reactants) at their active sites, resulting in an enzyme-substrate complex. This method can be summarized into three major steps:
Substrate Binding
The substrate interacts with the enzyme’s active site, which is usually a pocket or cleft in the enzyme’s structure. The active site’s unique amino acid configuration enables accurate substrate binding.
Transition State Formation
Once the substrate is bonded, the protein catalysts supports its conversion into products, stabilizing the transition state throughout the reaction.
Product Release
Following the reaction, the products are freed from the protein catalyst, which then returns to its original condition, ready to catalyze the next reaction.
Strategies for Purification of Enzymes
There are several ways for purifying protein catalyst from a mixture of other proteins. These strategies include the following:
Centrifugation
This approach divides components according to their size and density. Heavy particles (such as cell debris) settle to the bottom as the mixture is spun at high speeds, allowing the supernatant containing the soluble protein catalyst to be collected.
Precipitation
This method involves adding salts (such as ammonium sulfate) to selectively precipitate proteins, including protein catalyst, from the solution. By controlling the salt concentration, specific proteins can be separated based on their solubility.
Dialysis
Dialysis is used to remove small molecules and salts from a protein solution. The protein catalyst solution is placed in a dialysis bag or tube, which is then submerged in a buffer. Small molecules diffuse out, while larger protein catalystmolecules remain inside.
Chromatography
Various chromatography techniques can be utilized to separate enzymes based on different properties:
- Ion Exchange Chromatography: Separates protein catalyst based on charge.
- Affinity Chromatography: Uses specific binding interactions between the protein catalyst and a ligand attached to the chromatography medium.
- Gel Filtration Chromatography: Separates protein catalyst based on size, allowing smaller molecules to pass through while larger enzymes are retained.
Electrophoresis
This method separates protein catalysts based on their charge by applying an electric field to a gel. Different protein catalysts migrate at different rates, allowing for their separation and analysis.
Criteria of Purity
Specific Activity: This is defined as the quantity of product produced per unit time for each milligram of protein. Higher specific activity suggests higher protein catalyst purity.
Enzyme Activity: This evaluates the protein catalyst’s catalytic efficiency, which might be impacted by contaminants.
Homogeneity: Protein catalyst purity can be validated using procedures like SDS-PAGE (Sodium Dodecyl Sulfate Polyacrylamide Gel Electrophoresis), which separates proteins by size. A single band on the gel represents a homogenous enzyme preparation.
Isoelectric Focusing: This technique separates proteins based on their isoelectric point (pI), allowing for the identification of different protein species and confirming purity.
Amino Acid Analysis: Amino acid analysis can be used to determine the amino acid composition of a protein catalyst.
Molecular Weight Determination and Characterization of Enzymes
Gel filtration chromatography: Gel filtration chromatography can be used to separate proteins according to their size. The molecular weight of an protein catalysts can be calculated by measuring its elution volume from a gel filtration column.
SDS-PAGE: A technique that separates proteins by their molecular weight. The molecular weight of an protein catalysts can be calculated by comparing its migration distance in an SDS-PAGE gel to the migration distances of proteins with known molecular weight.
Mass Spectrometry: Provides accurate molecular weights and structural information.
Enzymes of Biological Importance
It plays crucial roles in various biological processes, including:
Digestion: Biocatalysts in the digestive system break down food into smaller molecules that can be absorbed into the bloodstream.
Respiration: Biocatalysts in the mitochondria catalyze the reactions that convert glucose into ATP, the energy currency of the cell.
Muscle contraction: Biocatalysts in muscle cells catalyze the reactions that allow muscles to contract.
DNA replication: Biocatalysts in the nucleus catalyze the reactions that replicate DNA.
Protein synthesis: Biocatalysts in the ribosomes catalyze the reactions that translate mRNA into proteins.
Applications of Enzymes
It is widely used in various fields:
Functional Group Interconversion
Biocatalysts can help in synthetic chemistry by allowing one functional group to be converted into another.
Retrosynthetic Biocatalysis
Biocatalysts are employed to break down complicated chemicals into simpler ones, providing information on synthesis routes.
Chemoenzymatic Synthesis of Natural Products
Combining chemical and enzymatic methods to synthesize complex natural products efficiently.
Enzymes from Extreme Thermophilic and Hyperthermophilic Microorganisms (Extremozymes)
These are microorganisms that live in severe temperatures, such as hot springs, hydrothermal vents, and volcanic settings. To survive and function in these harsh temperatures, they have developed unique adaptations, such as the development of heat-stable proteins known as extremozymes.
Key Characteristics of Extremozymes
Thermostability: These biocatalysts are highly stable at high temperatures, often retaining their activity at temperatures above 80°C.
Thermotolerance: They can withstand prolonged exposure to high temperatures without denaturation.
Optimal Activity at High Temperatures: Many extremozymes exhibit optimal activity at temperatures significantly higher than mesophilic biocatalysts.
Sources of Extremozymes
Extremozymes are primarily sourced from extremophilic microorganisms, including:
Hyperthermophilic Archaea: Such as Pyrococcus furiosus and Methanocaldococcus jannaschii, which produce biocatalysts that are stable and active at temperatures exceeding 100°C.
Thermophilic Bacteria: Such as Thermus aquaticus, which is known for its heat-stable DNA polymerase (Taq polymerase), widely used in PCR (Polymerase Chain Reaction).
Applications of Extremozymes
Extremozymes have numerous applications across various fields:
Pharmaceuticals: Their stability and activity at high temperatures make them useful in the production of pharmaceuticals that require rigorous processing conditions.
Biotechnology: Used in molecular biology techniques (e.g., PCR), where thermostable DNA polymerases are essential.
Industrial Processes: Employed in the food industry for processes like starch hydrolysis (amylases) and in biofuel production for breaking down biomass.
Bioremediation: Extremozymes can be used to degrade pollutants in extreme environments, such as hot springs or geothermal areas.
Conclusion
Enzymes are crucial biological catalysts that facilitate various chemical reactions in living organisms, playing key roles in processes such as digestion, respiration, and DNA replication. They can be classified into categories based on their functions and are sourced from plants, animals, and microorganisms. The purification of biocatalysts is essential for studying their properties and applications, utilizing techniques like centrifugation and chromatography to ensure their purity. Notably, extremozymes from extreme thermophilic and hyperthermophilic microorganisms exhibit remarkable stability and activity at high temperatures, making them valuable in industrial and biotechnological applications. Overall, biocatalysts enhance our understanding of biological processes while offering significant practical uses across multiple fields, including medicine and environmental science.
Frequently Asked Questions (FAQ)
What are enzymes?
Enzymes are biological catalysts that accelerate chemical processes in living organisms by decreasing the activation energy needed for the reaction to occur.
What factors affect enzyme activity?
Temperature, pH, substrate concentration, and the presence of inhibitors or activators can all have an effect on enzyme activity.
What are extremozymes?
Extremozymes are enzymes produced by microorganisms that live in extreme environments, such as high temperatures or pH levels. They are distinguished by their stability and activity in these severe environments.
