Deoxyribose-phosphate aldolase

Aldolases catalyze asymmetric aldol reactions via either Schiff base formation (type I aldolase) or activation by Zn2+ (type II aldolase) (Figure 1.16). The most common natural donors of aldoalses are dihydroxyacetone phosphate (DHAP), pyruvate/phosphoenolpyruvate (PEP), acetaldehyde and glycine (Figure 1.17) [71], When acetaldehyde is used as the donor, 2-deoxyribose-5-phosphate aldolases (DERAs) are able to catalyze a sequential aldol reaction to form 2,4-didexoyhexoses [72,73]. Aldolases have been used to synthesize a variety of carbohydrates and derivatives, such as azasugars, cyclitols and densely functionalized chiral linear or cyclic molecules [74,75]. 

The cholesterol-lowering drug atorvastatin, marketed as Lipitor, is an example where biocatalysis research has been applied extensively and is in industrial use. The enzyme 2-deoxyribose-5-phosphate aldolase (DERA) has been a target of directed evolution for the production of atorvastatin intermediates [8,9,71]. DeSantis and coworkers [8,9] used structure-based... 

DeSantis, G., Liu, J., Clark, D.R et al. (2003) Structure-based mutagenesis approaches toward expanding the substrate specificity of D-2-deoxyribose-5-phosphate aldolase. Bioorganic and Medicinal Chemistry, 11, 43-52. 

Figure 14.12 Asymmetric tandem aldol reaction using 2-deoxyribose-5-phosphate aldolase (DERA) and its application for production of Atorvastatin...

Gijsen, H.J.M. and Wong, C.-H. (1994) Unprecedented asymmetric aldol reactions with three aldehyde substrates catalyzed by 2-deoxyribose-5-phosphate aldolase. Journal of the American Chemical Society, 116 (18), 8422-8423. 

DEOXYRIBONUCLEASES 2 -Deoxyribonucleoside diphosphate, RIBONUCLEOTIDE REDUCTASE 2-DEOXYRIBOSE-5-PHOSPHATE ALDOLASE DEOXYTHYMIDINE KINASE DEP,... 

Concerning aldolases, the cloning of enzymes is becoming more and more common. Thus the bacterial fuculose-1-phosphate aldolase (EC 4.1.2.17) and 2-deoxyribose-5-phosphate aldolase (EC 4.1.2.4) have been recently overexpressed in E. coli and their synthetic use has been examined.115,116... 

To expand the substrate specificity and stereoselectivity of the aldolase DERA (2-deoxyribose-5-phosphate aldolase, E.C. 4.1.2.4), both site-specific mutagenesis and random mutagenesis have been investigated (DeSantis, 2003). The goal was to extend substrate specificity to the unnatural non-phosphorylated substrate, D-2-deoxyribose. 

An interesting enzyme-catalyzed three-component aldolization reaction has been described by Gijsen and Wong [18]. Here, acetaldeyde, 2-substituted acetaldehydes, and dihydroxyacetone phosphate react in the presence of the aldolases 2-deoxyribose-5-phosphate aldolase (DERA) and fructose 1,6-diphosphate aldolase (RAMA) forming the corresponding 5-deoxyketose derivatives (Scheme 9.9). 

The chiral 2,4-dideoxyhexose derivative required for the HMG CoA reductase inhibitors has also been prepared using 2-deoxyribose-5-phosphate aldolase (DERA).The reactions start with a stereospecific addition of acetaldehyde (44) (Fig. 18.14) to a substituted acetaldehyde to form a 3-hydroxyl-substituted butyraldehyde 45, which reacts subsequently with another acetaldehyde to form a 2,4-dideoxyhexose derivative 46. DERA has been expressed in Escherichia coli (Gijsen and Wong, 1995). 

Scheme 5.2. The four main groups of aldolase reactions classified by their donor substrate (1) Dihydroxyacetone phosphate (DHAP)- dependent aldolases, (2) phosphoenol pyruvate (PEP)-and pyruvate-dependent aldolases, (3) 2-deoxyribose-5-phosphate aldolase (DERA), a member of the acetaldehyde-dependent aldolases, and (4) glycine-dependent aldolases (GDA).

The enzyme DERA, 2-deoxyribose-5-phosphate aldolase (EC 4.1.2.4), is unique among the aldolases in that the donor is an aldehyde. In vivo it catalyzes the reversible aldol reaction of acetaldehyde and D-glyceraldehyde 3-phosphate, forming 2-deoxyribose 5-phosphate, with an equilibrium lying in the synthetic direction (Scheme 5.41). DERA, the only well-characterized member of this type I aldolase, has been isolated from both animal tissue and microorganisms.67... 

Machajewski TD, Wong CH. The catalytic asymmetric aldol reaction. Angew. Chem. Int. Ed. Engl. 2000 39(8) 1352—1375. Heine A, Luz JG, Wong CH, Wilson lA. Analysis of the class 1 aldolase binding site architecture based on tbe crystal stracture of 2-deoxyribose-5-phosphate aldolase at 0.99A resolution. J. Mol. Biol. 2004 343(4) 1019-1034. 

A procedure for large-scale production of 2-deoxy-5-thio-D-eryf/tro-pentose (O Scheme 15) has been developed. It uses a recombinant 2-deoxyribose-5-phosphate aldolase (DER A) from E. coli strain DH5a as the catalyst that combines acetaldehyde with racemic 3-thioglyceralde-hyde [98]. 

A different aldolase has been over-expressed in E. coli and used by Chi-Huey Wong in his synthesis of epothilones. It is 2-deoxyribose-5-phosphate aldolase (DERA) and the natural reaction is the condensation of acetaldehyde as enol with glyceraldehyde-3-phosphate 164 as electrophilic component to give an aldol product 165 that is trapped as a hemiacetal 166. 

Figure 3. Use of 2-deoxyribose-5-phosphate aldolase in the synthesis of fluorinated sugars, (a) Normal reaction, (b) synthesis of l-fluoro-4-hydroxy-5-methyl-hexan-2-one, (c) synthesis of 2,5-dideoxy-5-fluororibose...

This chapter describes recent developments in carbohydrate synthesis using microbial aldolases. Recombinant type II fructose-1,6> diphosphate aldolase and type I 2-deoxyribose-5-phosphate aldolase have been exploited for the synthesis of monosaccharides and analogs. A new procedure for the synthesis of dihydroxy acetone phosphate has been developed. Sialic acid aldolase has been used in conjunction with other enzyme-catalyzed modifications of monosaccharides for the synthesis of sialic acid-related sugars. 

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