Specialized Nucleotide Metabolism and its Molecular Role in Modulating Microbial Community Dynamics

With the support of the CLP program in the Division of Chemistry, Professor Crawford from Yale University is investigating the role of nucleotide inhibitors of protein translation that are encoded in microbiomes across diverse body sites. The microbiome of humans and other animals is now considered to be an organ-like system, digesting our food, regulating host behaviors, training the immune system, and producing diverse nutrients, signaling molecules, and/or toxins to effectively compete and survive in their local environment.

However, the molecular mechanisms and fundamental chemical understanding of how key members of the microbiome compete for effective colonization remain unknown. Colonization mechanisms underlie whether microbiomes across diverse animals are in balance. The proposed experiments will characterize the structures and functions of nucleotide translational inhibitors produced by microbiome members, how they are produced, and how they regulate community dynamics.

These efforts will allow graduate students to acquire training in specialized metabolism in the context of animal-microbe chemical interactions. This research is also integrated with an outreach program “Microbial Magic: Harnessing the Power of Bacteria” to introduce high school students to the science of the microbiome and microbial metabolism.

The Crawford laboratory discovered a wide family of mis-annotated tRNA synthetases in microbiomes that are dedicated to the synthesis of a new class of specialized nucleotides featuring a remarkable orthoester functionality. These metabolites, which very likely evolved from translational machinery, do not contribute to cellular growth, but rather were found to inhibit translation.

While nucleoside natural products are well known, the phosphate group on these functionalized nucleotides prevents passive cellular diffusion. This limitation demands specialized uptake mechanisms in recipient organisms and potentially allows for selective taxonomic targeting across diverse microbiomes. The proposed studies combine chemistry, biochemistry, and microbiology interdisciplinary approaches to provide new insights into this family of mis-annotated tRNA synthetases that have remained “hidden in plain sight” in genome databases until now.

Project goals include structural characterization of nucleotides from “orphan” pathways, biochemical characterization of the enzymes involved in nucleotide biosynthesis, and functional characterization of the molecules in in vitro translation inhibition and their regulatory effects on microbiome community structure. The information gained from this work will provide new molecular insights on how microbiome members employ specialized metabolism to regulate microbiome structure and function across diverse environments.

This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.