Structure, Function, and Stability of Enzymes Covalently Attached to Single-Walled Carbon Nanotubes

Abstract

We describe the structure, activity, and stability of enzymes covalently attached to single-walled carbon nanotubes (SWNTs). Conjugates of SWNTs with three functionally unrelated enzymes-horseradish peroxidase, subtilisin Carlsberg, and chicken egg white lysozyme-were found to be soluble in aqueous solutions. Furthermore, characterization of the secondary and tertiary structure of the immobilized proteins by circular dichroism and fluorescence spectroscopies, respectively, and determination of enzyme kinetics revealed that the enzymes retained a high fraction of their native structure and activity upon attachment to SWNTs. The SWNT-enzyme conjugates were also more stable in guanidine hydrochloride (GdnHCl) and at elevated temperatures relative to their solution counterparts. Thus, these protein conjugates represent novel preparations that possess the attributes of both soluble enzymes-high activity and low diffusional resistance-and immobilized enzymes-high stability-making them attractive choices for applications ranging from diagnostics and sensing to drug delivery.

Development of effective nanobiocatalytic systems through the immobilization of hydrolases on functionalized carbon-based nanomaterials

“…As it can be seen in Table 5, all hydrolases underwent some conformational changes upon immobilization on both nanomaterials, in agreement with what was observed for other supports (Asuri et al, 2007;Cang-Rong and Pastorin, 2009;Karajanagi et al, 2004;Tzialla et al, 2010). The a-helix content of all covalentlyimmobilized enzymes on both nanomaterials increased compared to that of free enzymes.…”

“…Immobilization yields up to 55% were achieved, which results in an enzyme loading of 1.65 mg of enzyme per mg of nanomaterial. This enzyme loading value is particularly high compared to the published values for other types of carbon-based nanomaterials (Asuri et al, 2007;Cang-Rong and Pastorin, 2009;Karajanagi et al, 2004). In previous work, we have shown that the immobilization yield could be improved by using a lower enzyme to nanomaterial weight ratio, albeit at the expense of the enzyme loading (Pavlidis et al, 2010a).…”

Cell-penetrating and cell-targeting peptides in drug delivery

“…Radio-frequency waves can be used in this case because of the particular electronic properties of carbon nanotubes and they are more attractive than the near-infrared laser heating method used in others strategies. Moreover, carbon nanotubes could be derivatized with various proteins to improve their specificity [110]. Indeed, it might be feasible to link these nanotubes to CPPs, to improve their cellular uptake, or to targetingpeptides to concentrate them at the tumor site.…”

A review on carbon nanotubes in an environmental protection and green engineering perspective

“…Nonetheless, the introduction of CNTs as bioelectrodes constitutes a major breakthrough in EFC. Other than mediating charge transfer, CNTs have manifested great efficiency in EFC by supplying a strong platform for enzyme immobilization (Fischback et al, 2006;Asuri et al, 2007;Li et al, 2008b;Zhao et al, 2009;Zhou et al, 2009). CNTs allowed enzyme molecules to be covalently attached onto their surface and promote high enzyme loading by permitting crosslinked enzyme aggregate coating (Fischback et al, 2006).…”

“…In addition, the nanoscale environment provided by CNTs could enhance enzyme stability. Enzymes covalently attached onto CNTs were found to have a high degree of stability due to their covalent linkage, which afforded significant resistance against denaturation (Govardhan, 1999;Asuri et al, 2007;Sheldon, 2007). In another point of view, the curvature of CNTs increases the distance between enzyme molecules, thereby reducing detrimental interaction between the enzymes and leading to an increased enzyme stability (Asuri et al, 2006).…”