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Stabilization of Immobilized Enzymes via the Chaperone-Like Activity of Mixed Lipid Bilayers

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posted on 16.05.2018, 00:00 by Andres F. Chaparro Sosa, Daniel F. Kienle, Rebecca M. Falatach, Jessica Flanagan, Joel L. Kaar, Daniel K. Schwartz
Biomimetic lipid bilayers represent intriguing materials for enzyme immobilization, which is critical for many biotechnological applications. Here, through the creation of mixed lipid bilayers, the retention of immobilized enzyme structures and catalytic activity are dramatically enhanced. The enhancement in the retention of enzyme structures, which correlated with an increase in enzyme activity, is observed using dynamic single-molecule (SM) fluorescence methods. The results of SM analysis specifically show that lipid bilayers composed of mixtures of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) and 1,2-dioleoyl-sn-glycero-3-phospho-(1′-rac-glycerol) (DOPG) stabilize the folded state of nitroreductase (NfsB), increasing the rate of refolding relative to unfolding of enzyme molecules on the bilayer surface. Remarkably, for optimal compositions with 15–50% DOPG, over 95% of NfsB remains folded while the activity of the enzyme is increased as much as 2 times over that in solution. Within this range of DOPG, the strength of the interaction of folded and unfolded NfsB with the bilayer surface was also significantly altered, which was evident by the change in the diffusion of folded and unfolded NfsB in the bilayer. Ultimately, these findings provide direct evidence for the chaperone-like activity of mixed DOPG/DOPC lipid bilayers, which can be controlled by tuning the fraction of DOPG in the bilayer.