Enzymology 1st Lecture PDF

# ENZYMES ## Definition - Enzymes are **biocatalysts** synthesized by living cells. - They are **protein** in nature, **colloidal** and **thermolabile** in character, and **specific** in action. - A **catalyst** is defined as a substance that increases the velocity or rate of a chemical reaction wi...

# ENZYMES ## Definition - Enzymes are **biocatalysts** synthesized by living cells. - They are **protein** in nature, **colloidal** and **thermolabile** in character, and **specific** in action. - A **catalyst** is defined as a substance that increases the velocity or rate of a chemical reaction without itself undergoing any change in the overall process. - **Friedrich Wilhelm Kuhne (1878)** - gave the name ENZYME (En = in; Zyme = yeast). - **James B. Sumner (1926)** - first isolated & crystallized urease from jack bean and identified protein. ## Nomenclature and Classification ### 1. Substrate Acted Upon by the Enzyme - Enzymes are named by adding the "ase" in the name of the substrate catalyzed. - Examples: - Carbohydrates - carbohydrases - Proteins - proteinases - Lipids - lipases - Nucleic acids - nucleases - Some names are more specific, like: - Maltase - acting upon maltose - Sucrase - upon sucrose - Urease - upon urea etc. ### 2. Type of Reaction Catalyzed - Enzymes are named by adding the "ase" in the name of the reaction. - Examples: - Hydrolases - catalyzing hydrolysis - Isomerases - isomerization - Oxidases - catalyzing oxidation - Dehydrogenases - catalyzing dehydrogenation - Transaminases - catalyzing transamination - Phosphorylases - catalyzing phosphorylation etc. ### 3. Substrate Acted Upon and Type of Reaction Catalyzed - Some enzyme names give clues about both the substrate utilized and the type of reaction catalyzed. - Examples: - Succinic dehydrogenase - catalyzes dehydrogenation of the substrate succinic acid. - L-glutamic dehydrogenase - catalyzes dehydrogenation reaction involving L-glutamic acid. ### 4. Substance That Is Synthesized - Some enzymes are named by adding the "ase" to the name of the substance synthesized. - Examples: - Rhodonase - forms rhodonate from hydrocyanic acid and sodium thiosulphate. - Fumarase - forms fumarate from L-malate. ### 5. Endoenzymes and Exoenzymes - Enzymes that act within the cells in which they are produced are called **intracellular enzymes** or **endoenzymes**. - Examples: plant enzymes and metabolic enzymes - Enzymes that are liberated by living cells and catalyze reactions outside the cell are called **extracellular enzymes** or **exoenzymes**. - Examples: bacterial enzymes, fungal enzymes, digestive tract enzymes ## International Union of Biochemistry (IUB) Nomenclature and Classification - The chemical reaction catalyzed is the specific property that distinguishes one enzyme from another. - In 1961, IUB used this criterion as a basis for the classification and naming of enzymes. - IUB classifies reactions and the enzymes catalyzing them into six major **classes**, each with four to thirteen **subclasses**. ### Enzyme Classes 1. Oxidoreductases 2. Transferases 3. Hydrolases 4. Lyases or Desmolases 5. Isomerases 6. Ligases or Synthetases ### 1. Oxidoreductases - Enzymes that catalyze oxidation-reduction reactions between two substrates. - General reaction: $AH_2 + B \longrightarrow A + BH_2$ - Groups present in the substrate: CH—OH, C=O, CH—CH, CH—NH₂, and CH=NH. - Examples: Alcohol dehydrogenase, Acyl-CoA dehydrogenase, Cytochrome oxidase etc. ### 2. Transferases - Enzymes that catalyze the transfer of a group, G (other than hydrogen) between two substrates, S and S'. - General reaction: $S-G + S' \longrightarrow S + S'-G$ - These enzymes catalyze the transfer of one-carbon groups, aldehydic or ketonic residues and acyl, glycosyl, alkyl, phosphorus, or sulfur-containing groups. - Examples: Acyltransferases, Glycosyltransferases, Hexokinase ### 3. Hydrolases - These enzymes catalyze the hydrolysis of their substrates by adding constituents of water across the bond they split. - General reaction: $A-B + H_2O \longrightarrow AH + BOH$ - The substrates include ester, glycosyl, ether, peptide, acid-anhydride, C-C, halide, and P-N bonds. - Examples: glucose-6-phosphatase, pepsin, trypsin, esterases, glycoside hydrolases ### 4. Lyases (Desmolases) - These enzymes catalyze the removal of groups from substrates by mechanisms other than hydrolysis, leaving double bonds. - General reaction: $ \begin{aligned} &X \\ &| \\ &C-C \\ &| \\ &Y \end{aligned} \longrightarrow \begin{aligned} &C=C + X-Y \end{aligned} $ - These include enzymes acting on C-C, C—O, C—N, C-S, and C-halide bonds. - Examples: Aldolase, Fumarase, Histidase etc. ### 5. Isomerases - These enzymes catalyze interconversion of optical, geometric, or positional isomers by intramolecular rearrangement of atoms or groups. - General reaction: $A \longrightarrow A'$ - Examples: Alanine racemase, Retinene isomerase, Glucosephosphate isomerase etc. ### 6. Ligases - These enzymes catalyze the linking together of two compounds by utilizing the energy made available due to simultaneous breaking of a pyrophosphate bond in ATP or a similar compound. - General reaction: $ \begin{aligned} &A + B \\ &\longrightarrow A-B \\ &ATP \\ &ADP+Pi \end{aligned} $ - This category includes enzymes catalyzing reactions forming C-O, C-S, C-N, and C-C bonds. - Examples: Acetyl-CoA synthetase, Glutamine synthetase etc. ## Enzyme Nomenclature - The Enzyme Commission (EC) system uses a four-digit code to categorize enzymes. - **First digit**: Type of general reaction (e.g., Hydrolase) - **Second digit**: Subclass of enzyme reaction (e.g., glycosidase) - **Third digit**: Sub-subclass of enzyme reaction (e.g., hydrolyze O glycosyl groups) - **Fourth digit**: Indicates specific enzyme (e.g., Alpha Amylase) - For example, EC 3.2.1.1 refers to alpha-amylase. ## Chemical Nature of Enzymes ### 1. Simple-Protein Enzymes - These enzymes contain simple proteins only. - Examples: urease, amylase, papain etc. ### 2. Complex-Protein Enzymes - These enzymes contain conjugated proteins, meaning they have a protein part called an **apoenzyme** and a nonprotein part called a **prosthetic group** associated with the protein unit. - The two parts constitute what is called a **holoenzyme**. - **Conjugated-protein enzyme** $\iff$ **Protein part** + **Prosthetic group** - **Holoenzyme** $\iff$ **Apoenzyme** + **Coenzyme** - The activity of an enzyme depends on the prosthetic group that is tightly associated with the apoenzyme. - Sometimes, the prosthetic group is loosely bound to the protein unit and can be separated by dialysis. This dialyzable prosthetic group is called a **coenzyme** (organic nature) or **cofactor** (inorganic nature). ## Coenzymes - A coenzyme is a non-protein, organic, low molecular weight, and dialysable substance associated with enzyme function. - Coenzymes are often regarded as **second substrates** or **co-substrates** because they have affinity with the enzyme comparable with that of the substrate. - Types: - B-complex vitamin coenzymes - Non B-complex vitamin coenzymes | Coenzyme | Abbreviation | Biochemical Functions | |---|---|---| | Adenosine triphosphate | ATP | Donates phosphate, adenosine, and adenosine monophosphate (AMP) moieties. | | Cytidine diphosphate | CDP | Required in phospholipid synthesis as a carrier of choline and ethanolamine. | | Uridine diphosphate | UDP | Carrier of monosaccharides (glucose, galactose), required for glycogen synthesis. | | S - Adenosylmethionine (active methionine) | SAM | Donates methyl group in biosynthetic reactions. | | Phosphoadenosine phosphosulfate (active sulfate) | PAPS | Donates sulfate for the synthesis of mucopolysaccharides. | ## Cofactors - A cofactor is a non-protein, inorganic, low molecular weight, and dialysable substance associated with enzyme function. - Most cofactors are metal ions. - **Metal activated enzymes:** In these enzymes, metals form a loose and easily dissociable complex. - Examples: ATPase (Mg²⁺ and Ca²⁺), Enolase (Mg²⁺) - **Metalloenzymes:** In these enzymes, metal ion is tightly bound to the enzyme and is not dissociated. - Examples: alcohol dehydrogenase, carbonic anhydrase, alkaline phosphatase, carboxypeptidase, and aldolase (contain zinc); Phenol oxidase (copper); Pyruvate oxidase (manganese); Xanthine oxidase (molybdenum); Cytochrome oxidase (iron and copper). ## Active Site - The active site (or active center) of an enzyme is the small region at which the substrate binds and participates in catalysis. **Salient features:** - The existence of the active site is due to the tertiary structure of the protein. - The active site is made up of amino acids that are far from each other in the linear sequence of amino acids. - Active sites are regarded as clefts, crevices, or pockets occupying a small region in a big enzyme molecule. - The active site is not rigid; it is flexible to promote specific substrate binding. - Enzymes are specific in their function due to the existence of active sites. - The active site possesses a **substrate binding site** and a **catalytic site**. - The coenzymes or cofactors on which some enzymes depend are present as part of the catalytic site. - The substrate binds at the active site by weak noncovalent bonds. - The commonly found amino acids at the active sites are serine (mostly found), aspartate, histidine, cysteine, lysine, arginine, glutamate, and tyrosine. - The substrate binds the enzyme (E) at the active site to form an enzyme-substrate complex (ES). The product (P) is released after catalysis, and the enzyme is available for reuse. - $E + S \rightleftharpoons ES \longrightarrow E + P$ ## Mode of Enzyme Action - Two theories have been put forth to explain the mechanism of enzyme-substrate complex formation: - 1. **Lock and key model/Fischer's template theory** - 2. **Induced fit theory/Koshland's model** ### 1. Lock and Key Model/Fischer's Template Theory - Proposed by Emil Fischer. - The first model proposed to explain an enzyme-catalyzed reaction. - According to this model, the structure or conformation of the enzyme is rigid. - The substrate fits to the binding site just as a key fits into a lock or a hand into a glove. - The active site of an enzyme is a rigid, pre-shaped template where only a specific substrate can bind. - This model was not accepted because: - It does not give any scope for the flexible nature of enzymes. - It fails to explain many facts of enzymatic reactions. - It does not explain the effect of allosteric modulators. ### 2. Induced Fit Theory/Koshland's Model - This model was proposed by Koshland. - The active site is not rigid or pre-shaped. - The interaction of the substrate with the enzyme induces a fit or a conformation change in the enzyme, resulting in the formation of a strong substrate binding site. - The appropriate amino acids of the enzyme are repositioned to form the active site and bring about catalysis. - This model was accepted because: - It has sufficient experimental evidence from X-ray diffraction studies. - It explains the action of allosteric modulators and competitive inhibition on enzymes.