BIOCHEMISTRY - Textbook - Ostapchenko L. I. - 2012

Chapter 7. ENZYMOLOGY

7.4. Enzyme Action Mechanism

7.4.3. Molecular Mechanisms of Enzymatic Catalysis

The Mechanisms of Enzymatic Catalysis are determined by The Role of the functional groups within the enzyme's Active Site in the chemical reaction that converts a substrate into a product. There are two primary mechanisms of enzymatic catalysis: Acid-Base Catalysis and Covalent Catalysis.

METABOLISM/2.html">THE CONCEPT OF acid-base catalysis explains enzymatic activity through the participation of acidic groups (proton Donors) and/or basic groups (proton acceptors) in the chemical reaction. Acid-base catalysis is a widely occurring phenomenon. The amino acid residues that make up the active site possess functional groups capable of exhibiting properties of both acids and bases.

The Amino Acids involved in acid-base catalysis primarily include Cys, Tyr, Ser, Lys, Glu, Asp, and His. In their protonated form, their side chains act as acids (proton donors), and in their deprotonated form, as bases (proton acceptors). This dual capability of the active site's functional groups makes Enzymes unique biological catalysts, unlike non-biological catalysts, which can exhibit either acidic or basic properties, but not both. An example of acid-base catalysis is Liver Alcohol dehydrogenase, which utilizes Zn²⁺ as a cofactor and an NAD⁺ molecule as a coenzyme to catalyze The oxidation of alcohol (see Fig. 7.13):

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Fig. 7.13. Mechanism of acid-base catalysis illustrated by liver alcohol dehydrogenase:

I - the ethyl alcohol molecule has a binding site that ensures hydrophobic interaction between the active site and the methyl group of the alcohol; II - the positively charged zinc atom facilitates the abstraction of a proton

from the alcohol group of ethanol, resulting in a negatively charged oxygen atom. The negative charge is redistributed between the oxygen atom and the adjacent hydrogen atom, which is then transferred as a hydride ion to the fourth carbon atom of the nicotinamide ring of the NAD⁺ coenzyme; III - this process yields the reduced form NADH and acetaldehyde

Covalent catalysis is based on the attack of nucleophilic (negatively charged) or electrophilic (positively charged) groups of the enzyme's active site by substrate molecules, resulting in The formation of a transient covalent bond between the substrate and a coenzyme or an amino acid side chain (typically one) within the active site.

The action of Serine proteases, such as Trypsin, Chymotrypsin, and Thrombin, provides a classic example of covalent catalysis, where a covalent bond is formed between the substrate and the serine amino acid residue in the enzyme's active site. The term "serine proteases" stems from the fact that a serine residue is an integral component of the active site in all these enzymes and participates directly in catalysis. Let us examine The Mechanism of covalent catalysis using chymotrypsin as an example, which catalyzes the Hydrolysis of peptide bonds during Protein Digestion in the duodenum. Chymotrypsin substrates are Peptides containing amino acids with aromatic and cyclic hydrophobic radicals (Phe, Tyr, Trp), indicating the involvement of hydrophobic forces in the Formation of the enzyme-substrate complex. The mechanism of covalent catalysis by chymotrypsin is illustrated in Fig. 7.14.

Fig. 7.14. Mechanism of covalent catalysis in the Active Site of chymotrypsin

The side chains of Asp₁₀₂, His₅₇, and Ser₁₉₅ are directly involved in the catalytic act. A nucleophilic attack on the substrate's peptide bond leads to its Cleavage, forming a covalently modified serine intermediate known as acyl-chymotrypsin. The other peptide fragment is released As a result of the rupture of the Hydrogen bond between the peptide fragment and His₅₇ in the active site of chymotrypsin. The final stage of peptide bond hydrolysis involves the deacylation of acyl-chymotrypsin in the presence of a Water molecule, releasing the second fragment of the hydrolyzed protein and regenerating the active enzyme.