18O-assisted dynamic metabolomics for individualized diagnostics and treatment of human diseases.

Technological innovations and translation of basic discoveries to clinical practice drive advances in medicine. Today's innovative technologies enable comprehensive screening of the genome, transcriptome, proteome, and metabolome. The detailed knowledge, converged in the integrated "omics" (genomics, transcriptomics, proteomics, and metabolomics), holds an immense potential for understanding mechanism of diseases, facilitating their early diagnostics, selecting personalized therapeutic strategies, and assessing their effectiveness.

Electron spray ionization mass spectrometry and 2D 31P NMR for monitoring 18O/16O isotope exchange and turnover rates of metabolic oligophosphates.

A new method was here developed for the determination of (18)O-labeling ratios in metabolic oligophosphates, such as ATP, at different phosphoryl moieties (α-, β-, and γ-ATP) using sensitive and rapid electrospray ionization mass spectrometry (ESI-MS). The ESI-MS-based method for monitoring of (18)O/(16)O exchange was validated with gas chromatography-mass spectrometry and 2D (31)P NMR correlation spectroscopy, the current standard methods in labeling studies. Significant correlation was found between isotopomer selective 2D (31)P NMR spectroscopy and isotopomer less selective ESI-MS method.

Dynamic phosphometabolomic profiling of human tissues and transgenic models by 18O-assisted ³¹P NMR and mass spectrometry.

Next-generation screening of disease-related metabolomic phenotypes requires monitoring of both metabolite levels and turnover rates. Stable isotope (18)O-assisted (31)P nuclear magnetic resonance (NMR) and mass spectrometry uniquely allows simultaneous measurement of phosphometabolite levels and turnover rates in tissue and blood samples. The (18)O labeling procedure is based on the incorporation of one (18)O into P(i) from [(18)O]H(2)O with each act of ATP hydrolysis and the distribution of (18)O-labeled phosphoryls among phosphate-carrying molecules.

(31)P NMR correlation maps of (18)O/ (16)O chemical shift isotopic effects for phosphometabolite labeling studies.

Intramolecular correlations among the (18)O-labels of metabolic oligophosphates, mapped by J-decoupled (31)P NMR 2D chemical shift correlation spectroscopy, impart stringent constraints to the (18)O-isotope distributions over the whole oligophosphate moiety. The multiple deduced correlations of isotopic labels enable determination of site-specific fractional isotope enrichments and unravel the isotopologue statistics. This approach ensures accurate determination of (18)O-labeling rates of phosphometabolites, critical in biochemical energy conversion and metabolic flux transmission.

Structural basis of ubiquitin recognition by translesion synthesis DNA polymerase ι.

Cells have evolved mutagenic bypass mechanisms that prevent stalling of the replication machinery at DNA lesions. This process, translesion DNA synthesis (TLS), involves switching from high-fidelity DNA polymerases to specialized DNA polymerases that replicate through a variety of DNA lesions. In eukaryotes, polymerase switching during TLS is regulated by the DNA damage-triggered monoubiquitylation of PCNA.