Reactive environmental toxicants, such as electrophiles and oxidants, can damage proteins. If this damage lowers a protein’s stability, then the protein can misfold and present a proteotoxic threat to the cell. Despite this risk, the protein targets of abundant toxicants are not generally known. To identify proteins that are destabilized by cellular toxicant exposures, we have developed a mass spectrometry-based Hsp40 affinity profiling assay. In this assay, a mutant DNAJB8 serves as a recognition element for misfolded cellular proteins, allowing them to be isolated and quantified. We used this assay to identify destabilized protein following exposure of HEK293T cells to three structurally similar chloroacetanilide herbicides: alachlor, propachlor, and acetochlor. While hundreds of proteins are targeted by these herbicides, and most of these targets harbor experimentally reactive cysteine residues, the profiles of these herbicides are distinct and include many proteins that lack general haloacetamide reactivity. These finding suggest that neither warhead reactivity nor protein reactivity on their own primarily drive targeting and destabilization. Two of the most selective propachlor targets are GAPDH and PARK7, both of which are strongly implicated in Parkinson’s Disease. We find that cellular activities of both proteins are inhibited by propachlor, with GAPDH inhibition driven by direct propachlor conjugation to C152. Destabilization of these proteins by propachlor might contribute to the strong association between occupational herbicide exposure and Parkinson’s Disease risk.

Cellular protein profiling to identify herbicide exposure targets

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