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Influence of Surface Charge of the Nanostructures on the Biocatalytic Activity Sirimuvva Tadepalli, Zheyu Wang, Keng-Ku Liu, Qisheng Jiang, Joseph M Slocik, Rajesh R. Naik, and Srikanth Singamaneni Langmuir, Just Accepted Manuscript • Publication Date (Web): 12 Jun 2017 Downloaded from http://pubs.acs.org on June 13, 2017
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Influence of Surface Charge of the Nanostructures on the Biocatalytic Activity Sirimuvva Tadepalli1, Zheyu Wang1, Keng-Ku Liu1, Qisheng Jiang1, Joseph Slocik2, Rajesh R. Naik2*, Srikanth Singamaneni1* 1
Institute of Material Science and Engineering and Department of Mechanical Engineering and Material Science, Washington University in St. Louis, St Louis, MO, 63130, USA. 2
Human Performance Wing, Wright-Patterson, Air Force Base, Dayton, OH
Abstract:
The physicochemical properties of abiotic nanostructures determine the structure and function of biological counterparts in biotic-abiotic nanohybrids. A comprehensive understanding of the interfacial interactions and the predictive capability of their structure and function is paramount for virtually all fields of bionanotechnology. In this study, using plasmonic nanostructures as a model abiotic system, we investigate the effect of the surface charge of nanostructures on the biocatalytic reaction kinetics of a bound enzyme. We found that the surface charge of nanostructures profoundly influences the structure, orientation and activity of the bound enzyme. Furthermore, the interactions of the enzyme with nanoparticles result in stable functional conjugates that retain their functionality at elevated temperatures, unlike their free counterparts that lose their secondary structure and biocatalytic activity.
*To whom correspondence should be addressed:
[email protected] (SS) and
[email protected] (RRN)
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Introduction
The design and synthesis of bionanoconjugates with precise and predictive control of their structure and function is important in a myriad of bionanotechnological applications, including biosensors, nanotheranostics, biocatalysis, artificial biosynthesis, bioinspired and bio-mediated energy harvesting devices, bioelectronic components, and molecular biology reagents.1-5 A fundamental understanding of the biomolecular structure at the interface and the function of bionanoconjugates has been elusive due to the complex interplay of various physicochemical properties (e.g., surface charge, charge distribution, surface functional groups, curvature) of the proteins and nanostructures.
6,7,8
The design of bionanoconjugates that retain the functionality
of the biomolecule (e.g., biorecognition, catalytic activity) is an indispensable tool in the progress of bionanotechnology.
It is known that the adsorption of proteins on nanoparticles causes conformational changes in the protein, leading to the exposure of new epitopes, altered function and avidity affects.9-11 The nature of protein adsorption determines the protein corona, which control the cellular interactions determining the fate of the nanoparticles.12-16 The formation and the nature of adsorption of proteins on nanoparticles depend on the nanoparticle size, shape and surface chemistry.17-22 In the design of bio-inorganic interfaces, it is important to have the enzymes in the right orientation in order to maximize electron transport.23-25 The orientation of the enzyme at the interface determines the relative position of the active site to the inorganic surface which determines the functionality of the enzyme.26 The biomolecular structure at this interface is critical in determining the functionality of the bionanoconjugates.
We hypothesize that the
surface chemistry of the nanostructure (gold nanoparticles with charged capping layer) modulates the biomolecular interactions at the interface, which govern the functionality of the bionanoconjugates. Interactions between the proteins and nanoparticles (electrostatic, van der
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Waals, hydrogen bonding, hydrophobic interactions) during and after adsorption alter the secondary structure of proteins and hence its function.27,28
The changes in the secondary
structure and the binding orientation of the enzymes on the nanoparticles determine the functionality of the bionanoconjugates.
We employed horseradish peroxidase (HRP) as the model enzyme to probe the influence of surface charge of the nanostructures on the biocatalytic activity of the bionanoconjugates. HRP is one of the most widely studied peroxidase that is extensively used in various bioanalytical applications.29,30
Apart from the well-recognized conformational changes, adsorption of
hemeproteins on nanostructures alters the availability of the distal side of the heme plane to the substrate during enzyme turnover. Hence, it is important to rationalize the biocatalytic activity of bionanoconjugates in terms of (i) the secondary structural changes due to the adsorption of the protein on nanoparticles and the accessibility of the heme binding site to the substrate in the bionanoconjugates. A deeper understanding of the structure and biophysicochemical properties of bionanoconjugates can lead to the rational design of bio-enabled nanozymes with synergistically enhanced biocatalytic properties.
Results and Discussion
Gold nanoparticles (AuNPs) were synthesized using a seed-mediated method (Figure 1(A), see experimental section for details).31 The diameter of the AuNPs was found to be 21.8 ± 0.9 nm from the TEM images. The controlled growth of nanoparticles from seeds through the slow addition of Au precursor into the growth solution resulted in a narrow size distribution of the nanoparticles (relative standard deviation
