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Genes encoding 2-deoxy-D-ribose-5-phosphate aldolase (DERA) homologues from two hyperthermophiles, the archaeon Pyrobaculum aerophilum and the bacterium Thermotoga maritima, were expressed individually in Escherichia coli, after which the structures and activities of the enzymes produced were characterized and compared with those of E. coli DERA. To our surprise, the two hyperthermophilic DERAs showed much greater catalysis of sequential aldol condensation using three acetaldehydes as substrates than the E. coli enzyme, even at a low temperature (25°C), although both enzymes showed much less 2-deoxy-D-ribose-5-phosphate synthetic activity. Both the enzymes were highly resistant to high concentrations of acetaldehyde and retained about 50% of their initial activities after a 20-h exposure to 300 mM acetaldehyde at 25°C, whereas the E. coli DERA was almost completely inactivated after a 2-h exposure under the same conditions. The structure of the P. aerophilum DERA was determined by X-ray crystallography to a resolution of 2.0 Å. The main chain coordinate of the P. aerophilum enzyme monomer was quite similar to those of the T. maritima and E. coli enzymes, whose crystal structures have already been solved. However, the quaternary structure of the hyperthermophilic enzymes was totally different from that of the E. coli DERA. The areas of the subunit-subunit interface in the dimer of the hyperthermophilic enzymes are much larger than that of the E. coli enzyme. This promotes the formation of the unique dimeric structure and strengthens the hydrophobic intersubunit interactions. These structural features are considered responsible for the extremely high stability of the hyperthermophilic DERAs.Using acetaldehyde and D-glyceraldehyde-3-phosphate as substrates, 2-deoxy-D-ribose-5-phosphate aldolase (DERA; EC 4.1.2.4) catalyzes a reversible aldol reaction that generates 2-deoxy-D-ribose-5-phosphate (DRP) (1,17). DERA is unique in that it catalyzes the aldol reaction between two aldehydes, which serve as both the aldol donor and the acceptor components. In addition, DERA is the only aldolase known to accept three aldehydes in a sequential and stereoselective manner during an aldol condensation reaction, which makes it a particularly interesting potential biocatalyst for synthetic organic chemistry. Gijsen and Wong (6) were the first to observe Escherichia coli DERA (DERA Eco ) catalyze a double aldol condensation of three acetaldehyde molecules (Fig. 1): the reaction started with a stereospecific addition of acetaldehyde to a substituted acetaldehyde to form 3-hydroxy-4-substituted butylaldehyde, which in turn reacted with a third acetaldehyde. After the second condensation, the product (compound 1) largely cyclized to form stable 2,4,6-trideoxy-D-erythro-hexapyranoside, which is a useful chiral synthon of hydroxymethylglutaryl-coenzyme A reductase inhibitors. This has prompted investigation of the feasibility of applying DERA Eco for the synthesis of cholesterol-lowering agents (5-7, 23). The practical application of th...

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Here we report the cellular arachidonate (AA)-releasing function of group IIF secretory phospholipase A 2 (sPLA 2 -IIF), a sPLA 2 enzyme uniquely containing a longer C-terminal extension. sPLA 2 -IIF increased spontaneous and stimulus-dependent release of AA, which was supplied to downstream cyclooxygenases and 5-lipoxygenase for eicosanoid production. sPLA 2 -IIF also enhanced interleukin 1-stimulated expression of cyclooxygenase-2 and microsomal prostaglandin E synthase. AA release by sPLA 2 -IIF was facilitated by oxidative modification of cellular membranes. Cellular actions of sPLA 2 -IIF occurred independently of the heparan sulfate proteoglycan glypican, which acts as a functional adaptor for other group II subfamily sPLA 2 s. Confocal microscopy revealed the location of sPLA 2 -IIF on the plasma membrane. The unique C-terminal extension was crucial for its plasma membrane localization and optimal cellular functions. sPLA 2 -IIF expression was increased in various tissues from lipopolysaccharidetreated mice and in ears of mice with experimental atopic dermatitis. In human rheumatoid arthritic joints, sPLA 2 -IIF was detected in synovial lining cells, capillary endothelial cells, and plasma cells. These results suggest that sPLA 2 -IIF is a potent regulator of AA metabolism and participates in the inflammatory process under certain conditions.

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