Biofunctionalization of α-zirconium phosphate nanosheets: toward rational control of enzyme loading, affinities, activities and structure retention.
Controlling the properties of enzymes bound to solid surfaces in a rational manner is a grand challenge. Here we show that preadsorption of cationized bovine serum albumin (cBSA) to α-Zr(IV) phosphate (α-ZrP) nanosheets promotes enzyme binding in a predictable manner, and surprisingly, the enzyme binding is linearly proportional to the number of residues present in the enzyme or its volume, providing a powerful, new predictable tool. The cBSA loaded α-ZrP (denoted as bZrP) was tested for the binding of pepsin, glucose oxidase (GOX), tyrosinase, catalase, myoglobin and laccase where the number of residues increased from the lowest value of ∼153 to the highest value of 2024. Loading depended linearly on the number of residues, rather than enzyme charge or its isoelectric point. No such correlation was seen for the binding of these enzymes to α-ZrP nanosheets without the preadsorption of cBSA, under similar conditions of pH and buffer. Enzyme binding to bZrP was supported by centrifugation studies, powder X-ray diffraction and scanning electron microscopy/energy-dispersive X-ray spectroscopy. All the bound enzymes retained their secondary structure and the extent of structure retention depended directly on the amount of cBSA preadsorbed on α-ZrP, prior to enzyme loading. Except for tyrosinase, all enzyme/bZrP biocatalysts retained their enzymatic activities nearly 90-100%, and biofunctionalization enhanced the loading, improved structure retention and supported higher enzymatic activities. This approach of using a chemically modified protein to serve as a glue, with a predictable affinity/loading of the enzymes, could be useful to rationally control enzyme binding for applications in advanced biocatalysis and biomedical applications.
Department of Chemistry, ‡Department of Molecular and Cell Biology, University of Connecticut , Storrs, Connecticut 06269, United States.
Inoka K
IK
Deshapriya
Christina S
CS
Kim
Marc J
MJ
Novak
Challa V
CV
Kumar
D000818:Animals; D055162:Biocatalysis; D002417:Cattle; D005949:Glucose Oxidase / Q000737:chemistry*; D008855:Microscopy, Electron, Scanning; D049329:Nanostructures / Q000737:chemistry*; D010434:Pepsin A / Q000737:chemistry; D011485:Protein Binding; D012710:Serum Albumin, Bovine / Q000737:chemistry*; D014961:X-Ray Diffraction; D015040:Zirconium / Q000737:chemistry*
C027006:zirconium phosphate; D012710:Serum Albumin, Bovine; D015040:Zirconium; D005949:Glucose Oxidase; D010434:Pepsin A