Recombinant Formaldehyde Dehydrogenase and Gene-Engineered Methylotrophic Yeasts as Bioanalitycal Instruments for Assay of Toxic Formaldehyde
Recombinant yeast clones, originated from the recipient Hansenula polymorpha strains NCYC 495 and CBS 4732, resistent to elevated concentrations of formaldehyde in a medium (up to 15–20 mM) and overproducing a homologous NAD- and glutathione-dependent form-aldehyde dehydrogenase, were constructed. Optimal cultivation conditions for the highest yield of the enzyme were established. A simple scheme for the isolation of formaldehyde dehydrogenase from the re-combinant strains was proposed, and some characteristics of the purified enzyme were studied. Enzymatic and biosensoric methods for formaldehyde assay based on the formaldehyde dehydrogenase and the constructed recombinant cells were developed. The reliability of the developed analytical approaches was tested on real samples of waste waters, pharmaceuticals, formaldehyde-containing industrial products, and vaccines. The comparison of formaldehyde content values obtained by the use of biosensors (enzyme and cells-based), enzymatic methods and two routinely used chemical ones (chromotropic acid and 3-methyl-2-benzothiazolinone hydrazone) showed a good correlation between these approaches.
KeywordsYeast Hansenula polymorpha (Pichia angusta) gene engineering formaldehyde dehydrogenase enzymatic assay biosensor formaldehyde
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