Ribose Utilization with an Excess of Mutarotase Causes Cell Death Due to Accumulation of Methylglyoxal

Abstract

Methylglyoxal (MG) is a highly reactive metabolic intermediate, presumably accumulated during uncontrolled carbohydrate metabolism. The major source of MG is dihydroxyacetone phosphate, which is catalyzed by MG synthase (the mgs product) in bacteria. We observed Escherichia coli cell death when the ribose transport system, consisting of the RbsDACBK proteins, was overproduced on multicopy plasmids. Almost 100% of cell death occurs a few hours after ribose addition (>10 mM), due to an accumulation of extracellular MG as detected by 1 H-nuclear magnetic resonance ( 1 H-NMR). Under lethal conditions, the concentration of MG produced in the medium reached approximately 1 mM after 4 h of ribose addition as measured by high-performance liquid chromatography. An excess of the protein RbsD, recently characterized as a mutarotase that catalyzes the conversion between the ␤-pyran and ␤-furan forms of ribose, was critical in accumulating the lethal level of MG, which was also shown to require ribokinase (RbsK). The intracellular level of ribose 5-phosphate increased with the presence of the protein RbsD, as determined by 31 P-NMR. As expected, a mutation in the methylglyoxal synthase gene (mgs) abolished the production of MG. These results indicate that the enhanced ribose uptake and incorporation lead to an accumulation of MG, perhaps occurring via the pentose-phosphate pathway and via glycolysis with the intermediates fructose 6-phosphate and glyceraldehyde 3-phosphate. It was also demonstrated that a small amount of MG is synthesized by monoamine oxidase.

Proteomic Analysis of Salmonella enterica Serovar Typhimurium Isolated from RAW 264.7 Macrophages

“…Interestingly, STM3119 is proposed to function as a monoamine oxidase (MaoA) that converts aminoacetone, which is the degradation product of L-threonine, to MG (Fig. 7) (51). As a well known fermentation product, D-lactate can also be used as a building block for synthesis of peptidoglycan (52).…”

Conversion of Methylglyoxal to Acetol by Escherichia coli Aldo-Keto Reductases

“…The CK281 [leu thr his thi rpsL lacY xyl ara tonA tsx ⌬rbs(D-R)::spc] strain harboring pJK5 (RbsD-overproducing plasmid derived from pBR322) and pJK10 (containing rbsACBK, derived from pACYC184) was used for nuclear magnetic resonance (NMR) analysis of metabolites from the intact cell. This CK281/pJK5/pJK10 strain exhibits an enhanced production of mutarotase, which results in methylglyoxal accumulation (20). Disruptions of the yqhE and yajO genes were created using a previously described method for chromosomal gene inactivation (9).…”

“…When we assessed their in vivo catalytic activities by creating double mutants, all of these genes except for yqhE exhibited further sensitivities to MG in a glyoxalase-deficient strain. The results imply that the glutathione-independent detoxification of MG can occur through multiple pathways, consisting of yafB, yqhE, yeaE, and yghZ genes, leading to the generation of acetol.Methylglyoxal (MG) is a widely occurring ketoaldehyde that is accumulated under physiological conditions with uncontrolled carbohydrate metabolism (1,11,20). MG synthesis is mediated by enzymes, including methylglyoxal synthase, cytochrome P450, and amine oxidase, which are involved in glycolytic bypass, acetone metabolism, and amino acid breakdown, respectively (8,18).…”

“…Methylglyoxal (MG) is a widely occurring ketoaldehyde that is accumulated under physiological conditions with uncontrolled carbohydrate metabolism (1,11,20). MG synthesis is mediated by enzymes, including methylglyoxal synthase, cytochrome P450, and amine oxidase, which are involved in glycolytic bypass, acetone metabolism, and amino acid breakdown, respectively (8,18).…”

Methylglyoxal detoxification by an aldo-keto reductase in the cyanobacterium Synechococcus sp. PCC 7002

“…3), suggesting that the gene product is involved in the protection of cells from toxic metabolites of glycerol. In E. coli, an uncontrolled carbon metabolism results in an increase in cellular methylglyoxal (Kim et al, 2004), and AKRs are required for its detoxification (Ko et al, 2005). The addition of glycerol to the growth medium induces a drastic increase in cellular methylglyoxal concentration in AKm1 (Fig.…”