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Chapter 3-1. Mechanism of Protein Degrading Enzymes

Recommended reading : 【Biology】 Chapter 3. Cells and Metabolism


Source : Sponsored by LG Chem Biotechnology Competition



Q. In general, the carbonyl group (C=O) in biochemical reactions is the atom attacked by nucleophiles. Therefore, the C=O double bond of the peptide bond (-CONH-) in a polypeptide chain can be hydrolyzed by water. This reaction is very slow, so it requires the help of enzymes, and proteases such as carboxypeptidase catalyze these reactions. For carboxypeptidase to have full activity, it requires a Zn(Ⅱ) metal ion. Based on this fact, answer the following questions.


⑴ Mechanism of Proteases


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Figure. 1. Mechanism of subtilisin, one of the proteases


Step 1. The carboxylate terminus of Asp32 nucleophilically attacks the imidazole ring of His64

Step 2. As a result, the conjugate base of the imidazole in His64 nucleophilically attacks the OH group of Ser221

Step 3. The alkoxide of Ser221 formed as a result nucleophilically attacks the carbonyl carbon of the protein

Step 4. The protein is broken down into carboxylic acid and amine by the base-catalyzed amide hydrolysis reaction

⑤ As implied by the term catalysis, the quantities of Asp32, His64, Ser221 do not change before and after the reaction


⑵ Types of Proteases

Pepsin : An endopeptidase that recognizes and cleaves the N-terminus of Phe, Trp, Tyr

Trypsin : An endopeptidase that recognizes and cleaves the C-terminus of Lys, Arg

Chymotrypsin : An endopeptidase that recognizes and cleaves the C-terminus of amino acids with a phenyl group (Phe, Trp, Tyr)

Carboxypeptidase : An exopeptidase that recognizes and cleaves the C-terminus of proteins

Subtilisin : A nonspecific protease obtained from Bacillus subtilis


⑶ Role of the Catalyst

① Answer why the reaction rate is faster when Zn(Ⅱ) metal ion is present at the active site of carboxypeptidase.


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Figure. 2. Mechanism of thermolysin, a type of carboxypeptidase


○ Carboxypeptidase, trypsin, and chymotrypsin recognize the C-terminus within the protein and cleave so that the carboxyl group is exposed

○ Basically, they use the acid-catalyzed hydrolysis reaction of amide


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Figure. 3. Acid-catalyzed hydrolysis reaction of amide


○ Zn(II) speeds up the reaction because it lowers the activation energy by electrostatically fixing the reaction environment

② Explain the role of histidine in the catalytic mechanism of chymotrypsin where serine nucleophilically attacks the peptide bond.

○ For the same reason as in ①. Histidine, along with lysine and arginine, is a positively charged amino acid that helps to fix the reaction environment like Zn(II).



Input: 2017.09.17 13:46

Modified: 2020.03.14 09:56

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