Identification of an evolutionary conserved structural loop that is required for the enzymatic and biological function of tryptophan 2,3-dioxygenase.

The enzyme TDO (tryptophan 2,3-dioxygenase; TDO-2 in Caenorhabditis elegans) is a potential therapeutic target to cancer but is also thought to regulate proteotoxic events seen in the progression of neurodegenerative diseases. To better understand its function and develop specific compounds that target TDO we need to understand the structure of this molecule. In C. elegans we compared multiple different CRISPR/Cas9-induced tdo-2 deletion mutants and identified a motif of three amino acids (PLD) that is required for the enzymatic conversion of tryptophan to N-formylkynurenine. Loss of TDO-2's enzymatic activity in PDL deletion mutants was accompanied by an increase in motility during aging and a prolonged lifespan, which is in line with the previously observed phenotypes induced by a knockdown of the full enzyme. Comparison of sequence structures suggests that blocking this motif might interfere with haem binding, which is essential for the enzyme's activity. The fact that these three residues are situated in an evolutionary conserved structural loop of the enzyme suggests that the findings can be translated to humans. The identification of this specific loop region in TDO-2-essential for its catalytic function-will aid in the design of novel inhibitors to treat diseases in which the TDO enzyme is overexpressed or hyperactive.

Delaying aging and the aging-associated decline in protein homeostasis by inhibition of tryptophan degradation.

Toxicity of aggregation-prone proteins is thought to play an important role in aging and age-related neurological diseases like Parkinson and Alzheimer's diseases. Here, we identify tryptophan 2,3-dioxygenase (tdo-2), the first enzyme in the kynurenine pathway of tryptophan degradation, as a metabolic regulator of age-related α-synuclein toxicity in a Caenorhabditis elegans model. Depletion of tdo-2 also suppresses toxicity of other heterologous aggregation-prone proteins, including amyloid-ß and polyglutamine proteins, and endogenous metastable proteins that are sensors of normal protein homeostasis. This finding suggests that tdo-2 functions as a general regulator of protein homeostasis. Analysis of metabolite levels in C. elegans strains with mutations in enzymes that act downstream of tdo-2 indicates that this suppression of toxicity is independent of downstream metabolites in the kynurenine pathway. Depletion of tdo-2 increases tryptophan levels, and feeding worms with extra L-tryptophan also suppresses toxicity, suggesting that tdo-2 regulates proteotoxicity through tryptophan. Depletion of tdo-2 extends lifespan in these worms. Together, these results implicate tdo-2 as a metabolic switch of age-related protein homeostasis and lifespan. With TDO and Indoleamine 2,3-dioxygenase as evolutionarily conserved human orthologs of TDO-2, intervening with tryptophan metabolism may offer avenues to reducing proteotoxicity in aging and age-related diseases.