Question

Proteins can be cleaved specifically at the amide bond on the carboxyl side of methionine residues by reaction with cyanogen bromide, \(\mathrm{BrC}=\mathrm{N}\). The reaction occurs in several steps: (a) The first step is a nucleophilic substitution reaction of the sulfur on the methionine side chain with BrCN to give a cyanosulfonium ion, \(\left[\mathrm{R}_{2} \mathrm{SCN}\right]^{+} .\) Show the structure of the product, and propose a mechanism for the reaction. (b) The second step is an internal \(S_{N}^{2}\) reaction, with the carbonyl oxygen of the methionine residue displacing the positively charged sulfur leaving group and forming a five-membered ring product. Show the structure of the product and the mechanism of its formation. (c) The third step is a hydrolysis reaction to split the peptide chain. The carboxyl group of the former methionine residue is now part of a lactone (cyclic ester) ring. Show the structure of the lactone product and the mechanism of its formation. (d) The final step is a hydrolysis of the lactone to give the product shown. Show the mechanism of the reaction.

Short answer

The peptide chain is cleaved at methionine via a series of nucleophilic substitutions and hydrolysis steps.

Step-by-step solution

Nucleophilic Substitution

In this step, the sulfur atom in the methionine residue of the protein acts as a nucleophile. It attacks the electrophilic carbon of the BrCN molecule, which has a partial positive charge due to the polarization of the C-Br bond. This results in the displacement of the bromide ion and the formation of a cyanosulfonium ion. The sulfur gains a positive charge because it is now bonded to an additional group (the cyanide group), forming \[\left[\text{{R}}_2 \text{{SCN}}\right]^+\] as the product.

Intramolecular SN2 Reaction

The carbonyl oxygen of the methionine residue forms a bond with the carbon attached to the sulfur, resulting in the displacement of the sulfur group, now a good leaving group due to its positive charge. This internal reaction creates a five-membered cyclic intermediate, which is stable due to its ring formation.

Lactone Formation via Hydrolysis

This step involves the formation of a lactone ring from the cyclic intermediate. Hydrolysis occurs, involving the nucleophilic attack of water on the carbonyl carbon of the amino acid, resulting in the opening of the ring. The leaving sulfur group facilitates the formation of a lactone product, where the carbonyl group is now part of a cyclic ester linking the peptide chain.

Hydrolysis of the Lactone

The lactone, which is a cyclic ester, undergoes hydrolysis typically under acidic or basic conditions. This process involves the addition of water, breaking the ester bond, resulting in the final production of an acid and an alcohol from the original lactone. This effectively cleaves the peptide at the methionine residue, completing the sequence of reactions.

Key concepts

Nucleophilic Substitution

In protein cleavage, nucleophilic substitution is an essential step. Here, the sulfur atom in the methionine side chain acts as a nucleophile. A nucleophile is a molecule or ion that donates an electron pair to form a chemical bond. The sulfur attacks the carbon in cyanogen bromide, which has a partial positive charge, making it an electrophile.
This interaction leads to the replacement of the bromine atom, forming a cyanosulfonium ion. The product, [ R_2 SCN ]^+, forms when sulfur bonds to the cyanide group.
Understanding nucleophilic substitution is vital here because it highlights how certain atoms or groups in a molecule can be selectively replaced to form new compounds.

• This reaction showcases the chemistry behind targeted protein modifications.
• It focuses on the ability of nucleophiles like sulfur to initiate changes in a molecule.

Cyanosulfonium Ion Formation

The formation of a cyanosulfonium ion is a crucial part of cleaving proteins. This step transforms the methionine residue's sulfur into a positively charged group.
First, the sulfur from the methionine attacks the carbon in BrCN, causing the bromine to leave as a bromide ion. This results in the sulfur gaining an extra bond and a positive charge, forming the cyanosulfonium ion.
The significant aspect of this reaction is its ability to stabilize the positively charged sulfur, which acts as a leaving group in subsequent reactions.

• The formation of the cyanosulfonium ion is pivotal for the internal reactions that follow, setting the stage for further chemical transformations.
• This step effectively primes the protein molecule for cleavage at specific sites.

Lactone Hydrolysis

Lactone hydrolysis involves breaking down a cyclic ester, formed in prior steps, into its constituent parts. This is a crucial phase in the protein cleavage process.
During hydrolysis, water acts as a nucleophile. It attacks the lactone's carbonyl carbon, opening the ring and breaking the ester bond.
This reaction ultimately converts the cyclic ester into potentially useful fragments, such as amino acids.

• Hydrolysis is often facilitated by acidic or basic conditions, which enhance the reactivity of water as a nucleophile.
• The breakdown of lactones to carboxylic acids and alcohols demonstrates the chemistry of ester bonds being disrupted, releasing simpler molecules.

Intramolecular SN2 Reaction

An intramolecular SN2 reaction is where a nucleophile within the same molecule displaces a leaving group. In cleaving proteins, this step follows cyanosulfonium ion formation.
Here, the carbonyl oxygen of methionine acts as a nucleophile, attacking the carbon bonded to the positively charged sulfur.
This results in the sulfur being displaced, as it is now a good leaving group, and forms a five-membered ring, ensuring stability.

• Intramolecular reactions are favored due to their efficiency in forming stable products, like cyclic structures.
• This step highlights the ability of molecules to undergo transformations based on internal reactivity.