Chemoproteomics Yields a Selective Molecular Host for Acetyl-CoA.

Chemoproteomic profiling is a powerful approach to define the selectivity of small molecules and endogenous metabolites with the human proteome. In addition to mechanistic studies, proteome specificity profiling also has the potential to identify new scaffolds for biomolecular sensing. Here, we report a chemoproteomics-inspired strategy for selective sensing of acetyl-CoA. First, we use chemoproteomic capture experiments to validate the N-terminal acetyltransferase NAA50 as a protein capable of differentiating acetyl-CoA and CoA. A Nanoluc-NAA50 fusion protein retains this specificity and can be used to generate a bioluminescence resonance energy transfer (BRET) signal in the presence of a CoA-linked fluorophore. This enables the development of a ligand displacement assay in which CoA metabolites are detected via their ability to bind the Nanoluc-NAA50 protein "host" and compete binding of the CoA-linked fluorophore "guest". We demonstrate that the specificity of ligand displacement reflects the molecular recognition of the NAA50 host, while the window of dynamic sensing can be controlled by tuning the binding affinity of the CoA-linked fluorophore guest. Finally, we show that the method's specificity for acetyl-CoA can be harnessed for gain-of-signal optical detection of enzyme activity and quantification of acetyl-CoA from cellular samples. Overall, our studies demonstrate the potential of harnessing insights from chemoproteomics for molecular sensing and provide a foundation for future applications in target engagement and selective metabolite detection.

Mycorrhizal type determines the magnitude and direction of root-induced changes in decomposition in a temperate forest.

Although it is increasingly being recognized that roots play a key role in soil carbon (C) dynamics, the magnitude and direction of these effects are unknown. Roots can accelerate soil C losses by provisioning microbes with energy to decompose organic matter or impede soil C losses by enhancing microbial competition for nutrients. We experimentally reduced belowground C supply to soils via tree girdling, and contrasted responses in control and girdled plots for three consecutive growing seasons. We hypothesized that decreases in belowground C supply would have stronger effects in plots dominated by ectomycorrhizal (ECM) trees rather than arbuscular mycorrhizal (AM) trees. In ECM-dominated plots, girdling decreased the activity of enzymes that break down soil organic matter (SOM) by c. 40%, indicating that, in control plots, C supply from ECM roots primes microbial decomposition. In AM-dominated plots, girdling had little effect on SOM-degrading enzymes, but increased the decomposition of AM leaf litter by c. 43%, suggesting that, in control plots, AM roots may intensify microbial competition for nutrients. Our findings indicate that root-induced changes in soil processes depend on forest composition, and that shifts in the distribution of AM and ECM trees owing to climate change may determine soil C gains and losses.