Benzaldehyde dehydrogenases

This pathway is supported by the demonstration of benzyl alcohol dehydrogenase, benzaldehyde dehydrogenase, benzoyl-CoA ligase, and benzoyl-CoA reductase activities in cell extracts (Biegert and Fuchs 1995). The benzyl alcohol dehydrogenase from benzyl alcohol-grown cells was similar in many of its properties to those from the aerobic bacteria Acinetobacter calcoaceticus and Pseudomonas putida (Biegert et al. 1995). 

Other approaches could use transcriptional regulators that directly bind the substrate or product of the reaction and activate the reporter gene. For instance, a mutant transcriptional activator from Pseudomonas putida, NahH, was used that can bind various benzoic acids to develop a screening/selection method to detect the action of benzaldehyde dehydrogenase [45]. A transcriptional regulator may need to be engineered to bind the desired compound before it can be used in such a manner [46]. 

Confirmation of this proposed pathway was established by Altenschmidt Fuchs (1991, 1992) in their study of the biochemistry of toluene decay by the denitrifying Pseudomonas sp. strain K172. These investigators confirmed the presence of benzyl alcohol dehydrogenase, benzaldehyde dehydrogenase, and benzoyl-CoA synthetase in cell-free extracts of this isolate. Further, [14C]benzyl... 

P. C. Babbitt, J. A. Gerlt, G. L. Kenyon, Identification and characterization of a mandelamide hydrolase and an NAD(P)+-dependent benzaldehyde dehydrogenase from Pseudomonas putida ATCC 12633,... 

Benzaldehyde dehydrogenase (BDH), a hypothetical enzyme, catalyzes the oxidation of benzaldehyde to benzoic acid. The enzyme has a flavin prosthetic group that accepts electrons from the substrate upon oxidation ... 

Benzaldehyde can be produced from benzoyl formate with whole cells of Pseudomonas putida ATCC 12633 as biocatalyst119 201 (Fig. 16.6-5). Alternatively, but less effectively, mandelic acid can be used as starting material. A pH of 5.4 was found to be optimal for benzaldehyde accumulation. At this proton concentration, partial inactivation of the benzaldehyde dehydrogenase isoenzymes and activation of the benzoyl formate decarboxylase are reported. Fed-batch cultivation prevented substrate inhibition. In situ product removal is necessary to prevent product inhibition. 

Scheme 3.11 Oxidation of toluene derivatives toward benzoic acid analogs (R = H, CHj) catalyzed by a gene cluster containing xylene-monooxygenase (XMO), benzylalcohol dehydrogenase (BADH), and benzaldehyde dehydrogenase (BZDH).

Another example on the use of metabolicaUy related enzymes was outbned by Zachariou [33] for the production of 3-carboxymuconate (3CM) from vanillin (Scheme 3.12). This study exploited the metabolic degradation of the cheap starting material vanillin by three different enzymes (4-hydroxy benzaldehyde dehydrogenase, vanillate monooxygenase, and protocatechuate 3,4-dioxygenase) to the monomer feedstock 3CM. After cloning the full pathway into E. coU, the authors were able to produce 3CM in very good overall HPLC yield (100% conversion) at concentrations up to 1 mM. 

---