Deciphering Reaction Mechanism with Intermediate Trapping Shuangyu Ma & Yiling Bi University of Utah

Identifying mechanism with intermediate trapping Overview: A reactive intermediate is a short-lived, high-energy, highly reactive molecule. When generated in a chemical reaction, it will quickly convert into a more stable molecule. When their existence is indicated, reactive intermediates can help explain how a chemical reaction takes place. Wiki Page: http://en.wikipedia.org/wiki/Reactive_intermediate Other References: Young, P. R.; Jencks, W. P. J. Am. Chem. Soc. 1977, 8238; Cordes, E.; Bull, H. Chem. Rev. 1974, 74, 581; Fife, T. Acc. Chem. Res. 1972, 78, 264.

Identifying mechanism with intermediate trapping Early Examples: hydrolysis of acetals, ketals and ortho esters: Wiki Page: http://en.wikipedia.org/wiki/Reactive_intermediate Other References: Young, P. R.; Jencks, W. P. J. Am. Chem. Soc. 1977, 8238; Cordes, E.; Bull, H. Chem. Rev. 1974, 74, 581; Fife, T. Acc. Chem. Res. 1972, 78, 264.

Identifying mechanism with intermediate trapping Trapping of the oxocarbonium ion intermediate by sulfite dianion: Results: achieved up to 97% trapping; rate constants for acid-catalyzed cleavage of ketals are independent of [SO32-] (Table 1). Conclusion: [SO32-] is not present in the TS of the RLS. Reaction proceeds through the oxocarbonium ion pathway. Wiki Page: http://en.wikipedia.org/wiki/Reactive_intermediate Other References: Young, P. R.; Jencks, W. P. J. Am. Chem. Soc. 1977, 8238; Cordes, E.; Bull, H. Chem. Rev. 1974, 74, 581; Fife, T. Acc. Chem. Res. 1972, 78, 264.

Identifying mechanism with intermediate trapping Early Examples: trapping of a single electron transfer intermediate. Proposed two pathways for the SN2 reaction : References: Haberfield, P. J. Am. Chem. Soc. 1995, 3314; Bank, S.; Noyd, D. A. J. Am. Chem. Soc. 1973, 95, 8203.

Identifying mechanism with intermediate trapping Early Examples: trapping of a single electron transfer intermediate. Using identity reaction and very large concentration of scavenger (as solvent). References: Haberfield, P. J. Am. Chem. Soc. 1995, 3314; Bank, S.; Noyd, D. A. J. Am. Chem. Soc. 1973, 95, 8203.

Application of intermediate trapping in understanding enzyme catalyzed complex reaction mechanism MoaA/MoaC catalyzed rearrangement reaction of GTP to cPMP: References: Mehta, A. P.; Abdelwahed, S. H.; Xu, H.; Begley, T. P. J. Am. Chem. Soc. 2014, 136, 10609; Mehta, A. P.; Abdelwahed, S. H.; Begley, T. P. J. Am. Chem. Soc. 2013, 135, 10883.

Application of intermediate trapping in understanding enzyme catalyzed complex reaction mechanism Trapping of the key intermediates using substrate analogues: References: Mehta, A. P.; Abdelwahed, S. H.; Xu, H.; Begley, T. P. J. Am. Chem. Soc. 2014, 136, 10609; Mehta, A. P.; Abdelwahed, S. H.; Begley, T. P. J. Am. Chem. Soc. 2013, 135, 10883.

Application of intermediate trapping in understanding enzyme catalyzed complex reaction mechanism References: Mehta, A. P.; Abdelwahed, S. H.; Xu, H.; Begley, T. P. J. Am. Chem. Soc. 2014, 136, 10609; Mehta, A. P.; Abdelwahed, S. H.; Begley, T. P. J. Am. Chem. Soc. 2013, 135, 10883.

Problems: 1. The scheme shown below is the proposed intramolecular anodic coupling amides and olefins. A second single-electron oxidation followed by solvent (MeOH) trapping of the subsequent cation would afford the final product 4. Based on the information, draw two possible pathways for the trapping of cation by the alcohol. (J. Org. Chem., 2014, 79, 379−391)

Problems: 2. Thymidylate synthases (TSases) catalyze the last step in the de novo biosynthesis of the DNA nucleotide dUMP to dTMP, using CH2H4 folate as a methylene donor. (J. Am. Chem. Soc., 2012, 134, 4442−4448) Figure 1 shows the total ion counts measured at various reaction times for substrate dUMP and product dTMP along with their sum (which represents the total amount of material due to these species). Please explain why intermediate accumulates according to data shown in Figure 1 and draw suggesting intermediate accumulation curve. Figure 1. Single-turnover FDTS reaction. Total ion counts for dUMP and dTMP. dUMP (purple◆) and dTMP (blue■). The sum of the counts for dUMP and dTMP (black▲) was not conserved during the reaction

Problems: (3) The figure shows (a) 14C-labeled dUMP or (b)14C-labeled CH2H4 folate. The labeled carbon is shown in red. In both cases, the 14C-containing trapped intermediate eluted at∼11 min, representing the same trapped species. According to the data shown below, please determine whether the trapped species already contains the methylene from the cofactor CH2H4 folate and justify your answer.

Solutions 1. The key part of the question is to come up with the equilibrium in the amidyl radical reactions. The radical products trapped by the alcohol are 3 and 4.

Solutions 2. In the figure, during 0.5 to 10 s, the sum of the ion counts for dUMP and dTMP was substantially less than those at the beginning and the end of the reaction. This observation suggests that a reaction intermediate accumulated during this time period. The suggesting intermediate accumulation curve is shown in red line.

Solutions 3. [11-14C] CH2H4 folate radiolabeled, a new radioactive peak had developed that had the same retention time as the peak observed when starting with [2-14C] dUMP. It means it is the same reactive intermediate. So it shows that the intermediate nucleotide being chemically trapped during the acid quenching had already undergone the condensation and that the carbon−carbon bond between the C5 of dUMP and the methylene had been formed prior to the formation of that intermediate.

Contributed by: Shuangyu Ma & Yiling Bi (Undergraduate Students) University of Utah 2014