Reagent targets methionines in proteins

Methods for chemically modifying proteins give researchers ways to attach probes, such as fluorescent dyes, or payloads, such as small-molecule drugs. Most chemical functionalization methods have focused on cysteine and lysine. There are far fewer methods that target methionine. Methionine is particularly attractive as a target for functionalization because it’s one of the least common amino acids, making modifications to it more selective.

Adding a new methionine modification method to the toolbox, Matthew J. Gaunt and coworkers at the University of Cambridge now report a way to alkylate surface methionines on proteins (Nature 2018, DOI: 10.1038/s41586-018-0608-y).

A previous method for functionalizing methionine, developed by Christopher J. Chang and F. Dean Toste of the University of California, Berkeley, uses redox chemistry to form stable sulfoximines. Gaunt and coworkers instead rely on a hypervalent iodine reagent as an electrophile to target the thioether in methionine’s side chain. The iodonium triflate they started with achieved only modest yields. “We had to engineer the reagent to make it more stable and more soluble in water,” Gaunt says. They swapped the triflate for tetrafluoroborate and added two fluorine atoms to the reagent’s aromatic ring. “Nobody in their right mind would think that this was going to be a reagent that reacts with proteins in water in a totally selective manner,” Gaunt says.

When the reagent reacts with methionine, it forms a sulfonium conjugate that includes a diazoester. The researchers then used the diazo group as a handle for the photocatalytic addition of other functional groups for further diversification of modifications. They made eight reagents with different diazoesters.

“You should be able to transfer any ester payload in this process,” Gaunt says. “As long as the payload you’re trying to transfer is fundamentally stable under the reaction conditions, this reaction should work.”

Adding a low concentration of thiourea speeds up the reaction, the researchers found. “Without the thiourea, the reaction still works. It takes maybe 30 minutes or so,” Gaunt says. “With thiourea, it’s almost instantaneous.” The thiourea isn’t acting as a denaturing agent, he says. He doesn’t yet know how the molecule speeds up the reaction, but his group is trying to elucidate its role.

The researchers used the reagent to functionalize a variety of polypeptides and proteins. Gaunt doesn’t have particular applications in mind but hopes it could help form antibody-drug conjugates or introduce imaging agents. “We’re a synthetic chemistry group, not a chemical biology group, and so we are trying to learn about how we could use this new chemistry in biological systems,” he says. “But hopefully it will be a general method that could be applied to whatever system anybody wanted to use.”

The work “offers an innovative and complementary approach to methionine bioconjugation that is chemically distinct from our oxidation-based method,” Berkeley’s Chang says. Chang is “very excited to see how these two approaches spawn more opportunities to study and augment protein function using this naturally occurring amino acid.”