Amide is one of the organic functional groups that can be found ubiquitously in the nature. It plays important role in living things as the main building blocks of proteins which involve with many functions, from catalysis to structural support. Besides that, amide functionality is featured in many natural products and drugs.
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| Examples of peptide-bearing compounds |
Since, it is quite common to find this functionality in natural products and synthetic compounds, a question is raised from this phenomenon. How trivial is it to make amide bond?
Answering this question, we need to see what amide functional group is. Amide functionality is one of the carboxylic acid derivatives, so when it is hydrolysed it will produce carboxylic acid and amine.
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| Amide hydrolysis |
Thus, it is quite sensible approach to say amide can be synthesised from those two functional groups. It turns out using both functionalities to amide is not as easy as it looks.
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| Chemical equilibria between acid-base reaction and amide formation |
The problem lies on the fundamental properties of carboxylic acid and amine. As its name suggested carboxylic acid is an acid, while amine is known as a base. Hence, it is inevitable that in aqueous condition acid base reaction will happen. Despite this problem, amide still can be made by ramping up the temperature to favour the formation of amide. Besides that, replacing the solvent with non-polar organic solvent can help the formation of amide. This is due the ions will not be solvated so it destabilise the formation of the acid-base product and favouring the amide.
As shown in the explanation above, amide can still be made from carboxylic acid and amine at high temperature and non-polar solvent. However, this statement still has a big question mark. In cellular level, the formation of amide bond (or peptide bond) happens in physiological temperature (27 °C) in aqueous environment; in these conditions the amide cannot be formed in a beaker. The obvious answer how proteins are made in the cells is they have 'the magic tools' of enzymes but if we examine the enzyme mechanisms how they make peptide bonds, it is surprisingly chemically sensible.
The way cells make peptide bonds from amino acids are helped by two major enzymes: aminoacyl-tRNA synthetase which joins amino acid with translator tRNA and the protein factory itself which is ribosome.
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| Aminoacyl-tRNA synthetase coupling mechanism |
The first step in protein biosynthesis is the coupling between zwitterionic amino acid with tRNA with the help from aminoacyl-tRNA synthetase. In the coupling reaction, it consumes one mole of ATP for every one mole amino acid. Because the amino acid is zwitterionic form, the carboxylate acts as nucleophile to attack the phosphate bond forming aminoacyl-AMP and pyrophosphate which then hydrolyse into two phosphate molecules. Then, it couple with tRNA producing aminoacyl-tRNA adduct. The highlight in this process is the formation of aminoacyl-AMP as the AMP itself in this adduct is a good leaving group and it facilitate the formation of ester functionality in the aminoacyl-tRNA adduct.
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| Ribosome and peptide formation mechanism at ribosome active site |
The next process, the tRNA travels to the protein factory ribosome where the peptide is made. Here, the peptide bond is made and the acid-base reaction is no longer a problem as the carboxylic acid has been transformed into ester functional group which is compatible with amine to form amide bond. In the ribosome itself, similar mechanism with the usual carbonyl reactions is happened. The amine as nucleophile attacks the carbonyl ester forming a tetrahedral intermediate which is stabilised by H-bonding interaction from an extended network of water and alcohol functional groups. Then, the tetrahedral intermediate collapses forming a peptide bond and releasing the tRNA in P-site which then move to E-site and displaced by newly formed peptide-tRNA adduct to continue the translation process.
The key message from the way cells forming peptide bond is transforming the carboxylic acid groups into a more compatible functionality which can react with basic amine functional group. This is the idea we can take in lab to make amide synthesis. The simple transformation would be the formation of acyl halide (usually acyl chloride) to react with amine forming amide functionality.
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| Amide formation from carboxylic acid via acyl chloride |
Another method is using coupling agent such as dicyclohexylcarbodiimide (DCC) and this coupling reaction is a common method in peptide-based drug syntheses.
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| Amide formation using DCC coupling agent |
The reaction proceeds via mechanism below.
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| The mechanism of amide formation using DCC coupling agent |
In this case, DCC is a better base than amine and it is protonated by the carboxylic acid forming carboxylate ions. As DCC is protonated, it becomes a better electrophile so it is now susceptible to nucleophilic attack by carboxylate forming the "activated acid". Then, the nuclephile amine can attack the carbonyl of the activated acid forming the usual tetrahedral intermediate and releasing the intended amide and DCU as side-product.
To conclude, the synthesis of amide is not as trivial as it may look. Taking the idea from the biosynthesis of peptide bond gives a better way to form amide bond in much milder condition and even producing higher yield. Moreover, biosynthesis-inspired synthetic methodologies has been a great interest in recent years to develop more efficient synthetic methodologies of certain target which might useful for daily life.
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