Biocatalytic reaction esterases

Formation of an amide bond (peptide bond) will take place if an amine and not an alcohol attacks the acyl enzyme. If an amino acid (acid protected) is used, reactions can be continued to form oligo peptides. If an ester is used the process will be a kinetically controlled aminolysis. If an amino acid (amino protected) is used it will be reversed hydrolysis and if it is a protected amide or peptide it will be transpeptidation. Both of the latter methods are thermodynamically controlled. However, synthesis of peptides using biocatalytic methods (esterase, lipase or protease) is only of limited importance for two reasons. Synthesis by either of the above mentioned biocatalytic methods will take place in low water media and low solubility of peptides with more than 2-3 amino acids limits their value. Secondly, there are well developed non-biocatalytic methods for peptide synthesis. For small quantities the automated Merrifield method works well. 

Hydrolases were in the first catalogue after the company was founded in 1950 but, not surprisingly, the chiral molecules originated mainly from the chiral pool. The first biocatalytic reactions were developed with kidney acylases and later with esterases and lipases, in the beginning mainly animal-derived biocatalysts [10], The set-up of in-house biocatalyst production from microbial and plant sources as well as the construction of a new biotechnology laboratory with ten fermenters of up to 300 L total volume, allow the development and production of improved biocatalysts and for them to be applied in the asymmetric synthesis of laboratory chemicals. There are today more than 100 biocatalytic processes in routine production and a project management team is handling custom biotransformations. 

The principal methods for the hydrolase-promoted synthesis of enantiomerically pure alcohols are depicted in Figure 6.44. Biocatalytic acylation and alcoholysis have been reviewed recently [116,117]. Lipases, esterases, and proteases catalyze these reactions, but CAL-B [118-120], CRL [121,122], and diverse lipase preparations from Pseudomonas species are common place. 

A number of major pharmaceutical companies have used biocatalytic approaches based on esterases and lipases lo prepare target drugs or intermediates [70,122-126]. Most of these approaches involve resolutions that start with a racemic ester or amide, and as such, yields of < 50% can only be realized. Recent examples of resolutions applied to pharmaceutical intermediates such as the paclitaxel (Taxol) side chain and A-(+)-BMY-14802, an antipsychotic agent, have been described by the Bristol-Myers group [70]. The following sections discuss selected examples of the use of esterases and lipases to hydrolyze prochiral or we o-substrates, where theoretical yields of 100% can be realized, followed by a brief discussion of dynamic kinetic resolution where reaction yields of 100% can also be achieved. 

Alternatively, surface hydrolysis of PET can be achieved by treatment with enzymes that introduce polar groups to the polymer surface. A number of hydrolytic enzymes, such as lipases, cutinases, and esterases, have shown potential for surface functionalization of PET [36, 99]. The biocatalytic method can be performed under mild reaction conditions avoiding the use of large amounts of chemicals and energy for the finishing and dyeing processes. The enzymatic modifications are specific and can be limited to the polymer surface. Consequently, the bulk properties and mechanical stability of the polymer are not compromised, and material savings and products of better quality or with new functionalities can be obtained. 

Since the beginning of enzyme catalysis in microemulsions in the late 1970s, several biocatalytic transformations of various hydrophilic and hydrophobic substrates have been demonstrated. Examples include reverse hydrolytic reactions such as peptide synthesis [44], synthesis of esters through esterification and transesterification reactions [42,45-48], resolution of racemic amino acids [49], oxidation and reduction of steroids and terpenes [50,51], electron-transfer reactions, [52], production of hydrogen [53], and synthesis of phenolic and aromatic amine polymers [54]. Isolated enzymes including various hydrolytic enzymes (proteases, lipases, esterases, glucosidases), oxidoreductases, as well as multienzyme systems [52], were anployed. 

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