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Abstract
BackgroundImmobilization of microbial cells is an important strategy for the efficient use of whole-cell catalysts because it simplifies product separation, enables the cell concentration to be increased, stabilizes enzymatic activity, and permits repeated or continuous biocatalyst use. However, conventional immobilization methods have practical limitations, such as limited mass transfer in the inner part of a gel, gel fragility, cell leakage from the support matrix, and adverse effects on cell viability and catalytic activity. We previously showed a new method for bacterial cell immobilization using AtaA, a member of the trimeric autotransporter adhesin family found in Acinetobacter sp. Tol 5. This approach is expected to solve the drawbacks of conventional immobilization methods. However, similar to all other immobilization methods, the use of support materials increases the cost of bioprocesses and subsequent waste materials.ResultsWe found that the stickiness of the AtaA molecule isolated from Tol 5 cells is drastically diminished at ionic strengths lower than 10 mM and that it cannot adhere in deionized water, which also inhibits cell adhesion mediated by AtaA. Cells immobilized on well plates and polyurethane foam in a salt solution were detached in deionized water by rinsing and shaking, respectively. The detached cells regained their adhesiveness in a salt solution and could rapidly be re-immobilized. The cells expressing the ataA gene maintained their adhesiveness throughout four repeated immobilization and detachment cycles and could be repeatedly immobilized to polyurethane foam by a 10-min shake in a flask. We also demonstrated that both bacterial cells and a support used in a reaction could be reused for a different type of reaction after detachment of the initially immobilized cells from the support and a subsequent immobilization step.ConclusionsWe invented a unique reversible immobilization method based on the salt-dependent adhesion of the AtaA molecule that allows us to reuse bacterial cells and supports by a simple manipulation involving a deionized water wash. This mitigates problems caused by the use of support materials and greatly helps to enhance the efficiency and productivity of microbial production processes.
Highlights
Immobilization of microbial cells is an important strategy for the efficient use of whole-cell catalysts because it simplifies product separation, enables the cell concentration to be increased, stabilizes enzymatic activity, and permits repeated or continuous biocatalyst use
Based on this molecular property of AtaA, we developed a unique method for the reversible immobilization of bacterial cells, which can solve the problems caused by the use of support materials
We found that the amounts of AtaA passenger domain (PSD) molecules adsorbed onto surfaces of hydrophobic PS and hydrophilic glass dropped sharply at ionic strengths lower than 10 mM, with the molecules hardly adhering in deionized water (dH2O), despite their high adhesiveness at higher ionic strengths (Fig. 1a)
Summary
Immobilization of microbial cells is an important strategy for the efficient use of whole-cell catalysts because it simplifies product separation, enables the cell concentration to be increased, stabilizes enzymatic activity, and permits repeated or continuous biocatalyst use. As for the downstream process, cell immobilization is important because it simplifies product separation, enables the cell concentration to be increased, stabilizes the enzymatic activity, and permits repetitive or continuous use of precious and expensive biocatalysts [12,13,14,15]. Conventional methods for cell immobilization are gel entrapment, covalent bonding to solid surfaces, cross-linkage, and physical adsorption [16, 17] These methods, have practical limitations, such as limited mass transfer in the inner part of a gel [18, 19], gel fragility, cell leakage from the support matrix, and adverse effects on cell viability and catalytic activity [12]. A way to minimize these drawbacks should be developed so as to, for example, reduce the amount of support materials, use inexpensive materials or waste materials, and reuse support materials
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