1: imidazole / dimethylformamide / 20 h / Ambient temperature 2: pyridine / 12 h 3: 94 percent / n-Bu4NF, AcOH / tetrahydrofuran / 18 h / Ambient temperature 4: 61 percent / diisopropylammoniumtetrazole / CH2Cl2 / 2 h 5: 62 percent / Et3N / acetonitrile / -43 deg C -> room temperature 6: Et3N, t-BuOOH / acetonitrile / 1 h / Ambient temperature 7: iPr2NH, (Ph3P)4Pd / acetonitrile / 0.5 h 8: NaOMe / methanol / 0.5 h 9: aq. NaOH / 0.17 h View Scheme

1: 61 percent / diisopropylammoniumtetrazole / CH2Cl2 / 2 h 2: 62 percent / Et3N / acetonitrile / -43 deg C -> room temperature 3: Et3N, t-BuOOH / acetonitrile / 1 h / Ambient temperature 4: iPr2NH, (Ph3P)4Pd / acetonitrile / 0.5 h 5: NaOMe / methanol / 0.5 h 6: aq. NaOH / 0.17 h View Scheme

CMP-Neu5Ac synthase; tri-(hydroxymethyl)-methylamine; magnesium chloride; DL-threo-1,4-dimercapto-2,3-butanediol T=37°C; 1.5 h; Enzymatic reaction;

1: E. coli sialic acid aldolase; water / 3 h / 37 °C / Enzymatic reaction 2: CMP-Neu5Ac synthase; tri-(hydroxymethyl)-methylamine; magnesium chloride; DL-threo-1,4-dimercapto-2,3-butanediol / 1.5 h / 37 °C / Enzymatic reaction View Scheme

recombinant Neisseria meningitidis CMP-sialic acid synthetase; recombinant sialic acid aldolase from Escherichia coli K12

sodium azide; acetate buffer T=37°C; also in D2O and in absence of azide; also in the presence of MeOH; solvent deuterium kinetic isotope effect; Rate constantProduct distributionMechanism;

in N,N-dimethyl-formamide T=105°C; 6 h;