Hydrogenase enzyme, stereospecific

Iron Sulfur Compounds. Many molecular compounds (18—20) are known in which iron is tetrahedraHy coordinated by a combination of thiolate and sulfide donors. Of the 10 or more stmcturaHy characterized classes of Fe—S compounds, the four shown in Figure 1 are known to occur in proteins. The mononuclear iron site REPLACE occurs in the one-iron bacterial electron-transfer protein mbredoxin. The [2Fe—2S] (10) and [4Fe—4S] (12) cubane stmctures are found in the 2-, 4-, and 8-iron ferredoxins, which are also electron-transfer proteins. The [3Fe—4S] voided cubane stmcture (11) has been found in some ferredoxins and in the inactive form of aconitase, the enzyme which catalyzes the stereospecific hydration—rehydration of citrate to isocitrate in the Krebs cycle. In addition, enzymes are known that contain either other types of iron sulfur clusters or iron sulfur clusters that include other metals. Examples include nitrogenase, which reduces N2 to NH at a MoFe Sg homocitrate cluster carbon monoxide dehydrogenase, which assembles acetyl-coenzyme A (acetyl-CoA) at a FeNiS site and hydrogenases, which catalyze the reversible reduction of protons to hydrogen gas. 

The pharmaceutical and fine chemical industry might use pure hydrogenase or partially purified enzyme preparations in bioconversion applications such as regio and stereoselective hydrogenation of target compounds (van Berkel-Arts et al. 1986). Enzymes are able to catalyse such stereospecific syntheses with ease. However, the cofactors for the NAD-dependent oxidoreductases are expensive. The pyridine nucleotide-dependent hydrogenases such as those from Ralstonia eutropha and hyperthermophilic archaea (Rakhely et al. 1999) make it possible to exploit H2 as a low-cost reductant. The use of inverted micelles in hydrophobic solvents, in which H2 is soluble, has advantages in that the enzymes appear to be stabilized. 

Stereospecific deuterations of 2-enoates (14,15,29,30,40) have to be carried out in H20 buffer with freeze dried cells (Table 5). Freeze drying under exclusion of oxygen leads to cells without loss of enzyme activity if again under anaerobic conditions 1 g wet packed cells is slowly stirred or shaken with 0,5 ml of a 10 pM solution of potassimn hexacyanoferrate-III for 20 minutes at 35 C. Under these conditions enoate reductase and probably also hydrogenase are transferred into the oxidized state. 

---