### Catalytic Mechanism of Thiolase in β-Oxidation of Saturated FatsThe enzyme thiolase plays a crucial role in one of the steps of β-oxidation, which is the metabolic process of breaking down saturated fats. This explanation provides a detailed overview of the mechanism as depicted in the schematic diagrams.#### Note:- **SCoA** refers to Acetyl-CoA.- Reaction: \( \text{RCOCH}_2\text{COSCoA} + \text{HSCoA} \rightarrow \text{H}_3\text{C}_2\text{OSCoA} + \text{RCOSCoA} \)#### Mechanism Steps:#### Diagram 1:- The substrate, \(\text{RCOCH}_2\text{COSCoA}\), enters the enzyme's active site.- The thiol group (\( \text{SCoA} \)) is bound to a carbonyl, interacting within the enzyme's catalytic site.#### Diagram 2:- A nucleophilic attack occurs on the carbonyl carbon of the substrate by the thiol group, forming a tetrahedral intermediate.- The intermediate rearranges to release Acetyl-CoA (\( \text{H}_3\text{C}_2\text{OSCoA} \)).#### Diagram 3:- The newly formed intermediate is further stabilized in the active site.- A second thiol group (\( \text{HSCoA} \)) attacks, leading to the formation of the thiolase product (\(\text{RCOSCoA}\)) and regenerating the enzyme for further catalysis.This sequence highlights the catalytic power of thiolase in cleaving the bond within the substrate, essential for β-oxidation pathways in metabolic processes.

Fatty acids are transported into the cell. The enzyme fatty acyl-CoA synthase(FACS) adds a CoA group to the fatty acid chain. This is called an acyl-CoA. The short-chain acyl-CoA chain can easily diffuse into the mitochondria, but the long-chain acyl-CoA chain is transported via the carnitine shuttle. The enzyme acyl CoA dehydrogenase oxidizes the acyl-CoA. There is a formation of a double bond between 2C and 3C. the end product is called trans-Δ2-enoyl-CoA. FAD is reduced to FADH2 in this step. Enoyl CoA hydratase adds a hydroxyl group to the 2C of trans-Δ2-enoyl-CoA to form L-β-hydroxyacyl CoA The 3-hydroxyacyl-CoA dehydrogenase catalyzes the oxidation of L-β-hydroxyacyl CoA to β-ketoacyl CoA and reduces NAD+ to NADH. β-ketoacyl CoA is cleaved by β-ketoacyl CoA thiolase (or simply thiolase) between 2C and 3C to produce acetyl-CoA. This acetyl CoA is made up of carbon 1C and 2C The new acyl-CoA chain which is now two carbons shorter will enter the second beta-oxidation cycle and with each cycle, it will be 2 carbons shorter.The reaction mechanism as described in the diagram: Deprotonated cysteine acts like a nucleophile and makes a nucleophilic attack on the C3 of β-ketoacyl-CoA. This results in the formation of a tetrahedral acyl-enzyme intermediate substitution product and the development of a negative charge on the carbonyl oxygen. Next, the negative charge on the oxyanion collapses. The bond between C2 and C3 is cleaved. The acyl intermediate (C3) is still attached to the cysteine group. The leaving group (C2) is protonated by the histidine. The acyl intermediate still attached to the cysteine residues is released after a CoA is attached to it.