Metabolism of carbohydrates and amino acids

Across all domains of life, organisms have evolved complex systems of interacting proteins with the task of mobilising sulfur atoms from the amino acid L-cysteine in the form of highly reactive persulfides (-S-S), which are then channeled to other proteins until they reach their ultimate destination — iron-sulfur (Fe-S) clusters, sulfur-containing vitamins, cofactors and lipids, or thiol-containing tRNA hypermodified ribonucleosides. Proper sulfur mobilisation and trafficking networks are therefore vital for life. In bacteria, two large systems exist that work either constitutively (ISC) or under oxidative stress (SUF) to ensure the correct routing of sulfur atoms across the metabolic network. In the model organism E. coli, a third system, CSD, has been characterised, although it remains poorly understood. Intriguingly, the CSD system is a minimal system with only two core genes (csdA and csdE) and an additional third gene (csdL/tcdA).

By using this minimalistic system as a model, we have determined the crystal structures of CsdA, CsdE and TcdA, the proteins encoded by the three CSD genes. Our ongoing work to characterise these proteins, how they interact with one another and how they function has led us to determine the structural and chemical bases for sulfur transfer across protein-protein surfaces. The findings can be transferred to the larger SUF system, since the PLP-dependent cysteine desulfurases in both systems, CsdA and SufS, are close homologues.

The connection with tRNA biology is made through TcdA, an E1-like enzyme capable of cyclising the N6-threonylcarbamoyl adenosine (t6A) present at the A37 position of the anti-codon stem loop (ASL) motif of tRNA(ANN) molecules. This modification, "cyclic t6A" (ct6A), has been recently discovered and has been proposed to be present across bacteria, protists, fungi and plants, where it would ensure the fidelity and efficiency of translation. We have investigated the structure of TcdA both free and associated with tRNA using a combination of X-ray crystallography/SAXS and other techniques, which represents one first step toward deciphering the biological role of this intriguing enzyme.

TcdA-tRNA complex