Research reveals new pathway in synthesis of thiostrepton, a powerful antibiotic

Utilizing X-ray crystallography, researchers at Penn State, MIT and cooperating institutions recently made a breakthrough in understanding the synthesis of thiostrepton, a powerful antibiotic with the potential to target even specific breast cancer cells. 

The drug has had limited use in veterinary medicine but has not been usable in humans because of its poor absorption. 

The new images, obtained with help of the Advanced Photon Source at Argonne National Laboratory and the Advanced Light Source at Lawrence Berkeley National Laboratory, have revealed the unusual chemical pathway followed in the first steps of thiostrepton biosynthesis. If these first steps can be mastered, there is hope that variants of the molecule can be produced with better curative properties.

The discovery was published Jan. 18 in the journal Nature Chemical Biology. 

Synthesis of thiostrepton begins with the essential amino acid tryptophan, known as the substrate. The tryptophan must then be modified by a process known as methylation – the replacement of a hydrogen atom by a methyl group (CH3), that is a carbon atom with three hydrogen atoms attached. 

In living organisms methylation is accomplished with the help of enzymes. Usually methylation occurs when an iron-sulfur cluster binds to the organic molecule S-adenosyl methionine (SAM-e), generating a so-called free radical that allows the reaction to progress. 

In the case, however, of thiostrepton biosynthesis, the protein involved known as TsrM, does not bind SAM-e to an iron-sulfur cluster but associates SAM-e directly with the tryptophan substrate used at the start the synthesis. In this first stage, the methyl group from the SAM-e is transferred to a portion of the TsrM protein known as cobalamin. Cobalamin is vitamin B-12. Then another SAM molecule helps to transfer the methyl group off cobalamin onto the tryptophan substrate.

Normally a methyl group would not be expected to move from cobalamin onto tryptophan. This is because cobalamin is the strongest nucleophile in nature, which means it holds onto things very tightly. But the X-ray crystallographic images revealed a new clue to how this might happen. The images showed that a key part, or residue, of another amino acid, arginine, was present below the cobalamin and apparently weakens the methyl-cobalamin bond. This allows the methyl group to move to tryptophan, regenerating free cobalamin for the next reaction.

Further research will examine ways to make use of the cobalamin displacement to produce antibiotics similar to thiostrepton that can be used in humans.