The Yi Tang group has discovered a new type of metal-containing enzyme from a natural fungus that can perform surprising halogenation reactions on organic molecules. The study, published in Nature, represents a four-year effort by graduate student Chen-Yu (Yorick) Chiang and research scientist Masao Ohashi, both in the Department of Chemical and Biomolecular Engineering.

Tang is the Parsons Family Foundation Professor in Chemical and Biomolecular Engineering, and also a Professor in Chemistry and Biochemistry, and Professor in Bioengineering at UCLA. This research represents a collaborative work from multiple research groups in the Department of Chemistry and Biochemistry, including those of Professors Joseph Loo, Ken Houk and Jose Rodriguez.  Additional collaborators include Professor Yisong (Alex) Guo at Carnegie Mellon University and Professor Shabnam Hematian from University of North Carolina at Greensboro.

The halogenation reaction, which involves the replacement of a carbon-hydrogen (C-H) bond with a carbon-halogen (C-X, X can be F, Cl, Br and I) bond in organic molecules, is a highly important synthetic transformation, particularly in the pharmaceutical industry.  Functionalization of molecules with halogen substitutents can dramatically improve the physical and pharmacological properties, as well as providing a reaction handle that can enable further modification of the molecules.  While many synthetic halogenation reagents have been developed, achieving selectivity, particularly introducing halogen at the least reactive position as this enzyme does, is remarkably difficult.

Nature has evolved enzymes known as halogenases, but  the known enzyme family uses iron as a reactive center to generate alkyl radicals to achieve halogenation.  However, the reaction scope is limited to substrates that can coordinate with the iron.  The new enzyme class involving copper will greatly expand the scope of enzymatic halogenations.

The Tang group focused on halogenated molecules produced by microorganisms in order to find new halogenases.  The group identified the molecule atpenin A5, a highly potent antifungal compound, featuring two carbon-chlorine bonds at unactivated carbon atoms.  By identifying enzymes in the biosynthetic pathway, a protein previously labeled as “domain of unknown function (DUF) 3328” was implicated in the halogenation reaction.  The authors worked with the Rodriguez lab and used AlphaFold protein structure prediction tool to identify a potential metal binding site that is different from known enzymes.  Different characterization techniques, including mass spectrometry (with Loo and graduate student Jessie Le), electron paramagnetic resonance (with Guo) and biochemical assays were used to show the DUF enzyme binds two copper atoms to catalyze the challenging halogenation reaction with high efficiency.  The use of a copper center in an enzyme to perform halogenation is unprecedented in Nature. 

With discovery of the copper-dependent enzyme, the Tang lab expanded enzymatic halogenation reaction that was not possible with the iron-based halogenases.  For example, the enzyme is able to perform iodination of an unactivated C-H bond that has not been demonstrated by enzymes.  This is due to the compatibility in electronic properties between copper and iodide, which is not feasible with iron.  Capitalizing on the unique property of the copper center, other useful functional groups such as thiocyanate and selenocyanate were also shown to be selectively incorporated into the substrate molecule.  Computational analysis by the Houk group, including postdoctoral scholar Panpan Chen and graduate student Qingyang Zhou, established a potential mechanism for the copper-dependent halogenation that will be explored in detail in the future.

“This discovery showed new enzymes with unprecedented properties and catalytic powers can be discovered from Nature,” Tang said of the discovery, “These enzymes perform reactions that are highly challenging by synthetic methods, and can therefore be useful tools for modifying or constructing molecules.”  It turns out this family of copper-dependent enzymes are widely found in Nature, most of which catalyze unknown reactions.  The Tang research team is now exploring functions of these previously cryptic enzymes to understand their potentials as biocatalysts.

For more information, contact Professor Yi Tang, yitang@g.ucla.edu