Regulate Binding Sites Of AuNPs With AChE By Combinatorial Nanoparticle Array Approach

Finding out the possible binding sites of nanoparticles (NPs) on proteins can benefit both the study of nanotoxicity and the preparation of functional bionanomaterials, helping to promote safer and more effective implementation of nanotechnology. The layer of adsorbed proteins on the surface of nanomaterials entering the biosystems is believed to serve as the biosignature of nanomaterials and impact their uptake, distribution, and excretion. The adsorption may also alter protein conformation and function, causing adverse health effects. By exploring where the NPs would bind on proteins, we can predict the biological consequence of protein adsorption and thus the potential toxicity. On the other hand, nanobio hybrid materials are prepared by attaching proteins to nanomaterials for energy production, sensing, separation,biomedical imaging, drug delivery, etc. The locations of nanomaterials on proteins could be strategically controlled to prevent any interference to protein function, to allow efficient energy or electron transfer between the proteins and the supporting materials, or to permit natural molecule recognition. Tactical coupling of nanomaterials to proteins calls for more knowledge about the dependence of binding epitope on nanomaterial properties.Gold nanoparticles (AuNP) can be easily synthesized with good stability, dispersion and biocompatibility. They has been most promising nanomaterials in biomedical field because they can be easily functionalized through Au-S covalent bond. As a biological catalyst composed of a protein molecule, Acetylcholinesterase(AChE) is generated in the liver and then secreted into the blood enzyme. To determine blood cholinesterase activity is not only an important factor of acute organophosphorus pesticide poisoning diagnosis and efficacy evaluation, but also as a clinical reflection of hepatic synthetic function. Studies have shown that the occurrence of many diseases are related to the rise of AChE activity, thus serum cholinesterase levels are important clinical index to determine the severity and recovery. To inhibit abnormal AChE is imperative. This project studied the binding sites of AuNP and AChE by various of analytical methods and obtained structure-activity relationship(SAR). It consists of the following two parts:The first part-Synthesis and characterization of different surface modified AuNP. Firstly, we synthesized ligand molecules using combinatorial chemistry for the later modification, and then characterized them with HPLC-MS and NMR. Subsequently, we synthesized surface-functionalize AuNPs and characterized them with TEM and zeta potential characterization. In the last, we quantitative the ligand number on AuNP.In the second part, we studied the interaction and possible binding sites between AChE and AuNP by studying the activity inhibition, fluorescence quenching and adsorption capacity. Combined with the experimental results in vivo and in vitro, we found some ligand modificated AuNP can bind on AChE catalytic sites specifically and there are some SAR between some modified AuNP and AChE catalytic sites.Comprehensive analysis of the above experimental results, we could get the following conclusions:Random surface modification of AuNP by combinatorial chemistry can change the NP surface properties. The binding sites are different with the different surface modification of AuNP. Most interactons are nonspecific while some ligand modified AuNP can specifically bind on AChE activity sites and induced more stability of AChE secondary structure. Combined with ligand structure and catalytic sites structure, we draw the conclusion that there may be some SAR between them. In the whole, surface molecular diversity by Combinatorial Nanoparticle Array Approach generate site-specific AuNP/enzyme recognition.