The biosynthesis of thiamine. Syntheses of [1,1,1,5-²H₄]-1-deoxy-D-threo-2-pentulose and incorporation of this sugar in biosynthesis of thiazole by Escherichia coli cells

Abstract

Non-growing, washed cells of Escherichia coli, depressed for the synthesis of thiamine, were incubated in the presence of [1,1,1 ,5-ZH4]-1 -deoxy-D-threo-2-pentulose (9) in a medium containing the pyrimidine moiety of thiamine, L-tyrosine, and glucose. The thiamine thus biosynthesized was extracted and cleaved to give 5-(2hydroxyethyl) -4-methylthiazole (H ET) which was examined as the trifluoroacetate derivative by electron-impact mass spectrometry. The dist:ibution of the label in the fragments indicated that the pentulose (9) was a precursor of the C,-chain of HET without C-C bond cleavage. Several routes to 1 -deoxypentuloses are described. Condensation of 2,4-O-benzylidene-D-[4-2H ,]threose (23) with trideuteriomethylmagnesium iodide gave the protected 1 -deoxypentitols ( 24) and ( 25). Brominolysis of the mixed dibutylstannylidenes then afforded [l,l,l ,5-2H4]-3,5-O-benzylidene-l -deoxy-D-threo-2-pentulose (26), which was converted into the free sugar (9) by acidic hydrolysis. 1 -Deoxy-D-erythro-2-pentulose was prepared in similar manner. Condensation of 2-( [2H3]methyl) -1,3-dithian with 2,3-O-isopropylidene-D-glyceraldehyde, followed by a C-3 epimerization step also led, after deprotection, to a mixture of [1,1,1 -2H3]-1 -deoxy-D-eryfhro-and [1,1,1 -2H3]-1 -deoxy-D-threo-2-pentulose, (5) and (6).

Suppression of insertions in the complex pdxJ operon of Escherichia coli K-12 by lon and other mutations

“…According to the scheme in Fig. 1, mutants deficient in this pathway branch should require vitamin B1 (thiamine), vitamin B2 (riboflavin), and PN (14,25,43). Future genetic searches will be aimed at finding mutants with multiple requirements besides PN and conditionally lethal mutants deficient in pyridoxine kinase activity.…”

Bioconversion of Lignocellulose: Inhibitors and Detoxifi cation

“…The same group improved upon the synthesis to deoxyxylulose and the [1-2 H 3 ]-isotopically-labeled derivative through condensation of aldehyde 2 with methylmagnesium iodide to provide the 2,4-O-benzylidene-D-threose intermediate 3. Following oxidation and deprotection, the free sugar 4 was obtained by brominolysis of the distannylidene derivative (Scheme 2) [21,22].…”

Isolation of a pdxJ point mutation that bypasses the requirement for the PdxH oxidase in pyridoxal 5' -phosphate coenzyme biosynthesis in Escherichia coli K-12

“…The pdxA and pdxB mutations block de novo synthesis of PNP (Fig. 1); however, PN taken up from the culture medium is phosphorylated to PNP by PL/PM/PN kinase in the B 6 vitamer salvage pathway (9,26,31). If the pdxJ(G194S) suppressor mutation provided an alternate oxidase activity in the pdxH null mutant, then PNP formed from exogenously added PN should support growth of the triple mutants.…”

“…The E. coli PdxH PNP/PMP oxidase plays roles in both the main de novo pathway of PLP biosynthesis ( Fig. 1) and the salvage pathway that interconverts the six B 6 vitamers, pyridoxal (PL), pyridoxamine (PM), pyridoxine (PN; vitamin B 6 ), PLP, PMP, and PNP ( Fig. 1) (9,18,19,26,31,40).…”