Secondary alcohol dehydrogenase

Figure 6 Enantioselective oxidation of secondary alcohols with secondary alcohol dehydrogenase (SADH), from a thermophihc bacterium (log E is used in...

A series of ethynyl ketones and ethynylketoesters were reduced enantioselectively to the corresponding nonracemic propargyl alcohols using a secondary alcohol dehydrogenase from... 

Yamamoto, H., Matsuyama, A. and Kobayashi, Y. (2002) Synthesis of ethyl (R)-4-chloro-3-hydroxybutanoate with recombinant Escherichia coli cells expressing (S)-specific secondary alcohol dehydrogenase. Bioscience Biotechnology and Biochemistry, 66 (2), 4814-83. 

Musa, M.M., Ziegelmann-Fjeld, K.I., Vieille, C. et al. (2007) Asymmetric reduction and oxidation of aromatic ketones and alcohols using W110A secondary alcohol dehydrogenase from Thermoanaerobacter ethanolicus. The Journal of Organic Chemistry, 72 (1), 30-34. 

Xerogel-encapsulated WHOA Secondary Alcohol Dehydrogenase from Thermoanaerohacter ethanolicus... 

Figure 9.5 Reduction of ketones with W110A secondary alcohol dehydrogenase...

Musa, M., Ziegelman-Fjeld, K., Vieille, C., Zeikus, J. and Phillips, R., Xerogel-encapsulated WHOA secondary alcohol dehydrogenase from Thermoanaerobacter ethanolicus performs asymmetric reduction of hydrophobic ketones in organic solvents. Angew. Chem. Int. Ed., 2007, 46, 3091-3094. 

Mori T, Sakimoto M et al (1998) Secondary alcohol dehydrogenase from a vinyl alcohol oligomer-degrading Geotrichum fermentans stabilization with Triton X-100 and activity toward polymers with polymerization degrees less than 20. World J Microbiol Biotechnoi 14 349-356... 

Pham, V.T., Phillips, R.S. and Ljungdahl, L.G. (1989) Temperature-dependent enantiospecificity of secondary alcohol dehydrogenase from Thermoanaerobacter ethanolicus. J. Am. Chem. Soc., 111, 1935-1936. 

The results of the temperature dependence of the reaction rates of the enantiomers of secondary alcohols with a secondary alcohol dehydrogenase (SADE1) from the thermophilic bacterium Thermoanaerobacter ethanolicus demonstrated a temperature-dependent reversal of stereospecificity (Pham, 1990) (Figure 5.16). At T < 26°C, (S)-2-butanol was a better substrate than (i )-2-butanol on the basis of kCSLt/KM values however, at T> 26°C, (R)-2-butanol was a better substrate than (S)-2-butanol. (S)-2-Pentanol was the preferred substrate at T < 60°C however, the data predict that (i )-2-pentanol would be preferred at T > 70°C. (S)-2-Elexanol was predicted to be the preferred enantiomer only at T > 240°C. Therefore, the concept of isoinversion temperature is as valid for enzyme reactions as for others only the range of catalytically accessible temperatures is smaller. 

C. Heiss, M. Laivenieks, G. J. Zeikus, and R. S. Phillips, The stereospecificity of secondary alcohol dehydrogenase from Thermoanaerobacter ethanolicus is partially determined by active site water,... 

RN Patel, CT Hou, AI Laskin, P Derelanko. Microbial production of methyl-ketones properties of purified yeast secondary alcohol dehydrogenase. J Appl Bio-chem 3 218-232, 1981. 

Many microorganisms possessing alcohol dehydrogenases that are capable of reducing ketones and diketones have been demonstrated to produce chiral alcohols. Examples of such enantioselective reductions have been reviewed on many occasions (3-8). The main advantage of a secondary alcohol dehydrogenase for the production of chiral alcohols... 

S. West, Production of chiral hydroxy groups by secondary alcohol dehydrogenase, Chim. Oggi., 9 43 (1990). 

Finally, researchers have also noted that the selection of non-aqueous solvents can affect enzyme activity in more subtle ways. Enzymes that bind with a dual pocket binding site (secondary alcohol dehydrogenases) possess enantiomeric conformations capable of binding (Cowan, 1997). It was observed that increasing the dielectric constant increased protein flexibility, which increased binding and catalysis. These conditions may also allow increased binding of the usually less favored enantiomer. 

From these results, the enzyme was found to be a NAD+-dependent (S)-specific secondary alcohol dehydrogenase. So far, only a few (S)-specific secondary alcohol... 

Cloning and Expression of a Gene Coding for a Secondary Alcohol Dehydrogenase from Candida parapsilosis IFO 1396 in Eschericha coli... 

Ex ample 1 The Thermoanaerobacter ethanolicus 39E adhB gene encoding the secondary alcohol dehydrogenase was overexpressed in Escherichia coli to form more than 10% to total protein11361. The recombinant enzyme was purified by heat treatment and precipitation with aqueous (NH4)2S04 and isolated in 67% yield. Enzymes with mutation(s) around the active site residues were also created to examine the catalytically important zinc binding motif in the proteins. 

The availability of sufficient quantities of enzymes for crystallization studies has led to the crystal structures been obtained for several dehydrogenases. For example, two tetrameric NADP+-dependent bacterial secondary alcohol dehydrogenases from the mesophilic bacterium Clostridium beijerinckii and the thermophilic bacterium Thermoanaerobium brockii have been crystallized in the apo- and the holo-enzyme forms, and their structures are available in the Protein Data Bank11451. The crystal structure of the alcohol dehydrogenase from horse liver is also available[40 21. 

Point mutation of enzymes has played an important role in determining those amino acid residues involved in catalytic activities. It has also been used to improve the enantioselectivity of dehydrogenases. For example, even a single point mutation of a secondary alcohol dehydrogenase from Thermoanaerobacter ethanolicus can change substantially the enantioselectivity for the reduction of 2-butanone and 2-pentanone as shown in Table 15-6 45l... 

Table 15-6. Control of enantioselectivity by a single mutation (serine-39 to threonine) of the secondary alcohol dehydrogenase from Thermoanaerobacter ethanolicus 5.

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