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Silica Gel for Enhanced Activity and Hypochlorite Protection of Cyanuric Acid Hydrolase in Recombinant Escherichia coli.

Radian A, Aukema KG, Aksan A, Wackett LP - MBio (2015)

Bottom Line: APTES coating or encapsulation of cells had two benefits: (i) overcoming diffusion limitations imposed by the cell wall and (ii) protecting against hypochlorite inactivation of CAH activity.APTES-encapsulated E. coli cells expressing CAH degraded cyanuric acid at high rates in the presence of 1 to 10 ppm hypochlorite, showing effectiveness under swimming pool conditions.Methods for promoting whole-cell biocatalysis are important in biotechnology, and the present work illustrates approaches to enhance rates and protect against an inhibitory substance.

View Article: PubMed Central - PubMed

Affiliation: BioTechnology Institute, University of Minnesota, St. Paul, Minnesota, USA Department of Mechanical Engineering, University of Minnesota, Minneapolis, Minnesota, USA.

No MeSH data available.


Related in: MedlinePlus

Scanning electron micrographs at two levels of magnification showing encapsulated E. coli cells in APTES gels with SNPs (APTES gel) (A) and E. coli cells coated by APTES (APTES-coated cells) (B). The scale bars of the top images are 2 µm, and the scale bars of the bottom images are 20 µm. Each white circle highlights a single cell to give another perspective of the size differences between the top and bottom images.
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fig2: Scanning electron micrographs at two levels of magnification showing encapsulated E. coli cells in APTES gels with SNPs (APTES gel) (A) and E. coli cells coated by APTES (APTES-coated cells) (B). The scale bars of the top images are 2 µm, and the scale bars of the bottom images are 20 µm. Each white circle highlights a single cell to give another perspective of the size differences between the top and bottom images.

Mentions: To overcome the diffusional barrier imposed by the cell membrane, a permeabilizing silica gel was developed. Previous studies have shown that appropriately positioned alkylamine functional groups disrupt Gram-negative cellular membranes, cause leakage of cytoplasmic materials, and render the bacteria nonviable (26–28). Since cyanuric acid-degrading enzymes use only water as a cosubstrate, cells expressing CAH can be nonviable and still show CYA degradation activity for weeks (25). In this context, a silica gel precursor with an amine functional group, 3-aminopropyltriethoxysilane (APTES), was chosen as the cross-linker for the silica gel matrix. Two treatments were applied to the cells to evaluate different levels of encapsulation in terms of diffusion and protection against hypochlorite inactivation. Figure 2A shows one approach in which APTES and silica nanoparticles (SNPs) were mixed to make a solid, continuous three-dimensional silica gel with the cells embedded within the gel. Figure 2B shows another process in which free cells were coated by particles formed by polymerization of hydrolyzed APTES that did not create a continuous gel but had a flake-like macrostructure. This was caused by cell clumping with a coated mesh-like appearance. The mesh would be expected to have a lower diffusional barrier than the gel, and it was expected that the amine groups on the APTES would render the membrane more porous.


Silica Gel for Enhanced Activity and Hypochlorite Protection of Cyanuric Acid Hydrolase in Recombinant Escherichia coli.

Radian A, Aukema KG, Aksan A, Wackett LP - MBio (2015)

Scanning electron micrographs at two levels of magnification showing encapsulated E. coli cells in APTES gels with SNPs (APTES gel) (A) and E. coli cells coated by APTES (APTES-coated cells) (B). The scale bars of the top images are 2 µm, and the scale bars of the bottom images are 20 µm. Each white circle highlights a single cell to give another perspective of the size differences between the top and bottom images.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4631802&req=5

fig2: Scanning electron micrographs at two levels of magnification showing encapsulated E. coli cells in APTES gels with SNPs (APTES gel) (A) and E. coli cells coated by APTES (APTES-coated cells) (B). The scale bars of the top images are 2 µm, and the scale bars of the bottom images are 20 µm. Each white circle highlights a single cell to give another perspective of the size differences between the top and bottom images.
Mentions: To overcome the diffusional barrier imposed by the cell membrane, a permeabilizing silica gel was developed. Previous studies have shown that appropriately positioned alkylamine functional groups disrupt Gram-negative cellular membranes, cause leakage of cytoplasmic materials, and render the bacteria nonviable (26–28). Since cyanuric acid-degrading enzymes use only water as a cosubstrate, cells expressing CAH can be nonviable and still show CYA degradation activity for weeks (25). In this context, a silica gel precursor with an amine functional group, 3-aminopropyltriethoxysilane (APTES), was chosen as the cross-linker for the silica gel matrix. Two treatments were applied to the cells to evaluate different levels of encapsulation in terms of diffusion and protection against hypochlorite inactivation. Figure 2A shows one approach in which APTES and silica nanoparticles (SNPs) were mixed to make a solid, continuous three-dimensional silica gel with the cells embedded within the gel. Figure 2B shows another process in which free cells were coated by particles formed by polymerization of hydrolyzed APTES that did not create a continuous gel but had a flake-like macrostructure. This was caused by cell clumping with a coated mesh-like appearance. The mesh would be expected to have a lower diffusional barrier than the gel, and it was expected that the amine groups on the APTES would render the membrane more porous.

Bottom Line: APTES coating or encapsulation of cells had two benefits: (i) overcoming diffusion limitations imposed by the cell wall and (ii) protecting against hypochlorite inactivation of CAH activity.APTES-encapsulated E. coli cells expressing CAH degraded cyanuric acid at high rates in the presence of 1 to 10 ppm hypochlorite, showing effectiveness under swimming pool conditions.Methods for promoting whole-cell biocatalysis are important in biotechnology, and the present work illustrates approaches to enhance rates and protect against an inhibitory substance.

View Article: PubMed Central - PubMed

Affiliation: BioTechnology Institute, University of Minnesota, St. Paul, Minnesota, USA Department of Mechanical Engineering, University of Minnesota, Minneapolis, Minnesota, USA.

No MeSH data available.


Related in: MedlinePlus