Identification of Chloroacetaldehyde Dehydrogenase Involved in 1,2-Dichloroethane Degradation

Abstract

The degradation of 1,2-dichloroethane and 2-chloroethanol by Xanthobacter autotrophicus GJ10 proceeds via chloroacetaldehyde, a reactive and potentially toxic intermediate. The organism produced at least three different aldehyde dehydrogenases, of which one is plasmid encoded. Two mutants of strain GJ1O, designated GJlOM30 and GJlOM41, could no longer grow on 2-chloroethanol and were found to lack the NAD-dependent aldehyde dehydrogenase that is the predominant protein in wild-type cells growing on 2-chloroethanol. Mutant GJlOM30, selected on the basis of its resistance to 1,2-dibromoethane, also had lost haloalkane dehalogenase activity and Hg2' resistance, indicating plasmid loss. From a gene bank of strain GJ10, different clones that complemented one of these mutants were isolated. In both transconjugants, the aldehyde dehydrogenase that was absent in the mutants was overexpressed. The enzyme was purified and was a tetrameric protein of 55-kDa subunits. The substrate range was rather broad, with the highest activity measured for acetaldehyde. The Km value for chloroacetaldehyde was 160 ,uM, higher than those for other aldehydes tested. It is concluded that the ability of GJ10 to grow with 2-chloroethanol is due to the high expression level of an aldehyde dehydrogenase with a rather low activity for chloroacetaldehyde.

Adaptations to a Loss-of-Function Mutation in the Betaproteobacterium Aromatoleum aromaticum: Recruitment of Alternative Enzymes for Anaerobic Phenylalanine Degradation

“…The majority of the recorded fragments correlated the enriched 55-kDa protein to the product of the aldB gene (ebA4625 [open reading frame designation]) from the genome of A. aromaticum. This protein has previously been annotated as an aldehyde dehydrogenase, showing 93% identity to an assumed acetaldehyde dehydrogenase of Thauera chlorobenzoica (33,34) or 80% amino acid identity to an NAD-dependent chloroacetaldehyde dehydrogenase of Xanthobacter autotrophicus (35). The aldB gene is part of an apparent operon with a gene coding for a Fe-dependent alcohol dehydrogenase (adhB, ebA4623) located immediately downstream.…”

Perspectives of genetically engineered microbes for groundwater bioremediation

“…Unwanted side reactions of chlorinated compounds can easily yield reactive products, and toxic effects of such reactive intermediates formed during metabolism of halogenated aliphatics are reported, also for the compounds discussed in this perspective. 40,65,74 The development of rapid tools for synthetic biology and genome engineering (e.g. DNA synthesis, combinatorial and multiplex methods for gene recombination and integration) will certainly accelerate the pace at which known bottlenecks can be solved, but the identication of these bottlenecks is still a tedious and time consuming task.…”

Elucidation of the Mechanisms and Environmental Relevance of cis-Dichloroethene and Vinyl Chloride Biodegradation

“…stopped by the addition of ZnSO 4 (30 mM) and were extracted with ethyl ether. Dichloroacetaldehyde dehydrogenase (van der Ploeg, Smidt et al, 1994) and chloroacetate dehalogenase (Janssen, Scheper et al, 1985) were measured by following the reduction of NAD to NADH at 365 nm.…”