Abstract
In this study, an efficient and generally applicable 2nd generation sol – gel entrapment method was developed for immobilization of yeastcells. Cells of Lodderomyces elongisporus, Candida norvegica, Debaryomyces fabryi, Pichia carsonii strains in admixture with hollow silica microspheres support were immobilized in sol – gel matrix obtained from polycondensation of tetraethoxysilane. As biocatalysts in theselective acyloin condensation of benzaldehyde catalyzed by pyruvate decarboxylase of the yeast, the novel immobilized whole-cell preparations were compared to other states of the cells such as freshly harvested wet cell paste, lyophilized cells and sol – gel entrapped preparations without hollow silica microspheres support. Reusability and storability studies designated this novel 2nd generation sol – gel method as a promising alternative for solid formulation of whole-cells bypassing expensive and difficult downstream steps while providing easy-to-handle and stable biocatalysts with long-term preservation of the biocatalytic activity.
Highlights
The biocatalytic systems based on whole-cells are blooming recently, due to the continuously growing interest on synthetic biology, protein engineering and recombinant techniques providing novel cells with extremely widespread biocatalytic activity of various origin [1, 2]
2.2.4 Immobilization of cells by the 2nd generation sol–gel method using hollow silica microspheres as support In case of 2nd generation sol – gel entrapment, a suspension was prepared by addition of MAT540 support (150 mg) to freshly harvested (0.5 g) yeast cells resuspended in phosphate buffer (3 mL, 0.1 M, pH 7.5)
2.2.7 Effect of co-factors and buffer systems on acyloin condensation of benzaldehyde 1 catalyzed by sol – gel entrapped yeast cells The acyloin condensation of benzaldehyde 1 catalyzed by Lodderomyces elongisporus yeast cells entrapped in 2nd generation sol – gel systems were carried out under similar conditions as described in Subsection 2.2.6 and in the presence of thiamine pyrophosphate (TPP: 10 n/n % and MgCl2: n/n %)
Summary
The biocatalytic systems based on whole-cells are blooming recently, due to the continuously growing interest on synthetic biology, protein engineering and recombinant techniques providing novel cells with extremely widespread biocatalytic activity of various origin [1, 2]. Whole-cell biocatalysts have many advantages compared to the isolated enzymatic systems, especially the avoidance of expensive and time-consuming downstream processes including enzyme purification represents significant benefits. The whole-cells suffer a few drawbacks such as low tolerance of organic solvents, difficult co-factor regeneration and sometimes lower activities and side reactions [5]. While yeast cells can express diverse enzymatic activities, they are easy-to produce and handle, stable and commonly non-pathogenic. Their redox, C-C coupling or hydrolytic enzyme sets get significant attention in several syntheses of chiral intermediates or compounds [6,7,8,9,10]. Amongst the yeast catalyzed biotransformations, the carbon-carbon condensation is one of the most important reaction to produce optically
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