382547 Confinement Protection Effects of Mesoporous Silica, Inspired By Chaperonin Complexes

Wednesday, November 19, 2014: 4:55 PM
International 5 (Marriott Marquis Atlanta)
Michele Lynch and Marc-Olivier Coppens, Chemical Engineering, University College London, London, United Kingdom

With the increasing use of biocatalytic enzymes across many fields of industrial manufacturing, more research must be done to stabilise enzymes and thereby increase their utility. The most common challenges in biocatalysis include unsuitable pH, high temperature, biological contaminants, and the presence of organic solvents [1,2]. Immobilisation onto nanostructured materials can combat the inherent instability of enzymes, particularly in the harsh environments encountered in biocatalysis.

Mesoporous silica SBA-15 shows great potential for applications in enzyme immobilisation. SBA-15 is typically formed as rod-shaped microparticles with cylindrical nanopores arrayed hexagonally along their axis. These pores simulate the environment found in the GroEL/ES complex, a pair of chaperone proteins found in bacterial cells that assist in the folding of new proteins [3]. The pores of SBA-15 have a hydrophilic core, similar to the GroEL/ES complex, and exhibit similar electrostatics with respect to protein adsorption. This study investigates the denaturation protection conferred to protein adsorbed to the internal surface of SBA-15 from exposure to proteases and extreme pH.

The model biocatalytic enzymes myoglobin and lysozyme were chosen for this study because of their well-characterised structure and simple activity assays. The SBA-15 used in this study is synthesised at a target pore diameter just larger than that of the enzymes, to lead to tight confinement upon adsorption [4, 5]. The SBA-15 is characterised using scanning electron microscopy (SEM), nitrogen sorption, and small angle x-ray scattering (SAXS). To investigate the protective effects of SBA-15, the enzymes are adsorbed to SBA-15 and these composites are exposed to a range of detrimental pHs. Other confined samples are additionally exposed to protease. Myoglobin activity is tested via peroxidase activity assay, which is measured with UV-Vis spectroscopy. Lysozyme activity is tested via chitinase activity assay, which is measured with fluorescence spectroscopy.

Our results suggest that confinement in tightly-fitting SBA-15 protects the enzymes from both acidic denaturation and proteolysis. More work must be done to investigate further protective effects against high temperature and organic solvents, and further studies will be required to generalise SBA-15’s protective effects to enzymes with more useful industrial applications.

References

[1] Sheldon & van Pelt Chem. Soc. Rev. 42, 6223–35 (2013)

[2] Tran & Balkus ACS Catal. 1, 956–968 (2011)

[3] Mayhew et al. Nature 379, 420–426  (1996)

[4] Sang & Coppens Phys. Chem. Chem. Phys. 13, 6689–6698 (2011)

[5] Sang et al. Langmuir 27, 13828–37 (2011)


Extended Abstract: File Not Uploaded