What is chemical biology? Native Chemical Ligation!

Disclaimer: I am not an expert. In fact, this series of blog posts is as informative  to me as it is to you. Probably even more so. My views and the views of people interviewed for this blog do not, in fact, reflect what exactly “chemical biology” is, but only a snapshot.  Please direct any comments or suggestions below!

The two subunits were synthesized separately

HIV-1 Protease, one of the first proteins synthesized via chemical ligation (PDB 3CDR)

On the last “What is Chemical Biology” post, several readers had comments that Chemical Biology was nothing more than a relabeled version of Pharmacology.  To be honest, they do have a point.  A lot of Chemical Biology that I have investigated so far has been drug discovery, which falls squarely in the realm of Pharmacology.  To that end, I have decided to find something that fits in this field that is more “pure science.”  Enter Native Chemical Ligation.

Native Chemical Ligation was born of the effort of the total synthesis of proteins.  Solid phase peptide synthesis (SPPS), as most of you probably know, is limited to around 50 residues as the peptide begins to form secondary/tertiary structure around  the resin bead it is attached to.  This secondary structure could reduce the (already not so great) efficiency of the coupling reactions.  In addition, by this point in a peptide synthesis, the overall yield is very low as well.  This results in some very major problems in the total synthesis of proteins.  How did bio-organic chemists solve this problem?  You can probably guess the answer by now.

Native Chemical ligation is born from Chemical Ligation, a method to connect two synthesized peptides together without the use of protecting groups on each and every side chain.  Chemical ligation used chemistries involving the formation of thioester or thioether bonds among other linkages.  While Chemical ligation resulted in the total synthesis of several peptides, there are several disadvantages to this technique, including the replacement of a nitrogen with a sulfur atom.  This is the equivalent of placing a soccer ball in the place of a tennis ball.  Naturally, this can result in problems in protein folding.  Native Chemical Ligation is a method by which a native peptide bond can be “installed” into a synthetic peptide structure.

Look!  Modification!

Native Chemical Ligation Mechanism (Credit: American Physiological Society)

The chemistry is surprisingly simple.  A S to N acetyl transfer upon ligation of the two peptide fragments results in a native peptide bond and an easily purified thioester side product.  This reaction is shown in the figure.  Native chemical ligation can also be used to synthesize glycerolized peptides at specific sites.  This was described in an article out of Nature.  Overall, a pretty cool mechanism for synthesizing whole proteins, and making slight alternations without using recombinational techniques.

Author: Sidechain Bob

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