PMC

A proposed mechanism for thiazole biosynthesis in eukaryotes.

Identification of Thi4 bound small molecules by HPLC (a). Analysis of the HPLC purified peak A : by UV-Vis spectroscopy (b), by 1H NMR spectroscopy (c) and by negative mode ESI-MS (d).

Active site structure of Thi4 (PDB code: 2GJC): (a) The structure of Compound 2 and its interactions with the enzyme. (b) Electron density confirming the thiazole carboxylic acid moiety of 2. The density is a composite omit map contoured at the 1σ level.

Confirmation of the structure 3 by its enzymatic conversion to thiamin phosphate 5: (a) Scheme showing the reactions performed. (b) HPLC analysis of the cleavage reaction of 2 to 3 and AMP: The blue trace shows the control reaction where no pyrophosphatase has been added to 2; The purple trace shows the cleavage reaction of 2 by the pyrophosphatase. (c) HPLC detection of the thiochrome phosphate 6 produced from 2: blue trace shows the control reaction with no HMP-PP added; Purple trace shows the actual coupling reaction.