Evolution of fungal tuberculosis necrotizing toxin (TNT) domain-containing enzymes reveals divergent adaptations to enhance NAD cleavage.

Tuberculosis necrotizing toxin (TNT) is a protein domain discovered on the outer membrane of Mycobacterium tuberculosis (Mtb), and the fungal pathogen Aspergillus fumigatus. TNT domains have pure NAD(P) hydrolytic activity, setting them apart from other NAD-cleaving domains such as ADP-ribosyl cyclase and Toll/interleukin-1 receptor homology (TIR) domains which form a wider set of products. Importantly, the Mtb TNT domain has been shown to be involved in immune evasion via depletion of the intracellular NAD pool of macrophages.

Novel Calcium-Binding Motif Stabilizes and Increases the Activity of Ecto-NADase.

Nicotinamide adenine dinucleotide (NAD) is an essential molecule in all kingdoms of life, mediating energy metabolism and cellular signaling. Recently, a new class of highly active fungal surface NADases was discovered. The enzyme from the opportunistic human pathogen was thoroughly characterized.

Identification of structural determinants of nicotinamide phosphoribosyl transferase (NAMPT) activity and substrate selectivity.

NAD homeostasis in mammals requires the salvage of nicotinamide (Nam), which is cleaved from NAD by sirtuins, PARPs, and other NAD-dependent signaling enzymes. Nam phosphoribosyltransferase (NAMPT) catalyzes the rate-limiting step in vitamin B3 salvage, whereby Nam reacts with phosphoribosyl pyrophosphate (PRPP) to form nicotinamide mononucleotide. NAMPT has a high affinity towards Nam, which is further enhanced by autophosphorylation of His247.

The integration of AlphaFold-predicted and crystal structures of human -3-hydroxy-l-proline dehydratase reveals a regulatory catalytic mechanism.

Computational methods for protein structure prediction have made significant strides forward, as evidenced by the last development of the neural network AlphaFold, which outperformed the CASP14 competitors by consistently predicting the structure of target proteins. Here we show an integrated structural investigation that combines the AlphaFold and crystal structures of human -3-Hydroxy-l-proline dehydratase, an enzyme involved in hydroxyproline catabolism and whose structure had never been reported before, identifying a structural element, absent in the AlphaFold model but present in the crystal structure, that was subsequently proved to be functionally relevant. Although the AlphaFold model lacked information on protein oligomerization, the native dimer was reconstructed using template-based and computational approaches.

The balance between NAD biosynthesis and consumption in ageing.

Nicotinamide adenine dinucleotide (NAD) is a vital coenzyme in redox reactions. NAD is also important in cellular signalling as it is consumed by PARPs, SARM1, sirtuins and CD38. Cellular NAD levels regulate several essential processes including DNA repair, immune cell function, senescence, and chromatin remodelling.

Structure of Thermococcus litoralis Δ-pyrroline-2-carboxylate reductase in complex with NADH and L-proline.

L-Hydroxyproline (L-Hyp) is a nonstandard amino acid that is present in certain proteins, in some antibiotics and in the cell-wall components of plants. L-Hyp is the product of the post-translational modification of protein prolines by prolyl hydroxylase enzymes, and the isomers trans-3-hydroxy-L-proline (T3LHyp) and trans-4-hydroxy-L-proline (T4LHyp) are major components of mammalian collagen. T4LHyp follows two distinct degradation pathways in bacteria and mammals, while T3LHyp is metabolized by a two-step metabolic pathway that is conserved in bacteria and mammals, which involves a T3LHyp dehydratase and a Δ-pyrroline-2-carboxylate (Pyr2C) reductase.