Construction Of Supramolecular Artifical Nanoenzyme With Hydrolytic Activity

Enzymes catalyze almost all chemical reactions with stereo- selectivities and specificities under mild conditions. Nature has created a lot of hydrolytic enzymes.They have the capavity to catalyse the hydrolysis of phosphate, protein and nucleic acid etc.They are very important for livings. Unfortunately, scientists can not fully understand the structures of nucleophosphatase as well as its catalysis mechanism in vivo at the present time. Fabrication of nucleophosphatase models offers an ideal alternative for elucidating the origin of substrate binding and catalysis of enzyme.Nanoparticles behavior their material properties significantly different from those of their bulk counterparts, because of the confinement and quantum effects. Their technological importance stems from their novel and useful structural, optical, and functional properties. In this regard, gold nanoparticles have emerged as prototypical models of metal nanoparticles with wide-ranging applications.We exploited the well known ability of thiols to bind to gold nanoparticles and prepared multivalent, functional and yet fully soluble systems anchored on a support-monolayer protected gold nanoparticles. This paper has done on preparation of gold nanoparticles and modified gold nanoparticles,as template mimic artifical metallic hydrolytic enzyme. The cleavage of the phosphate bond of phosphodiesters as a model of a RNase use this artifical metallic hydrolytic enzyme.1. The preparation of ligand and substrate HPNP. We designed and synthesized N1,N1-bis(2-aminoethyl)-N2-dodecyl- ethane-1.2-diamine. 1H NMR,IR and MALDI-TOF MS spectra were recorded. HPNP was prepared according to literature procedures.Its structure was indicated by 1H NMR spectroscopy.2.The preparation of water soluble functional gold nanoparticles. We used the method reported by Mathias Brust to prepare dodecanethiolate-functionalized gold nanoparticles. UV absorption spectroscopy and IR spectrum indicated that we had synthesized dodecanethiolate-functiona-lized gold nanoparticles.The dodecanethiolate-functionalized gold nanoparticles was dissolved in chloroform and added to the mixture of CTAB, Zn(Ⅱ) or Cu(Ⅱ)-bis (aminoethyl)dodecane aminoethyl amine.The chloroform was removed by vacuumto finish the encapsulation meanwhile the temperature was 40-45℃. A brown solution was finally obtained.We use dialytic method to purify Cu(Ⅱ) and Zn(Ⅱ)complexes of supramolecolar artifical nanoenzyme.3. The cleavage of the phosphate bond of phosphodiesters (HPNP) as a model of a RNase use this artifical metallic hydrolytic enzyme. Most of these enzymes require for their activity at least two metal ions that act cooperatively. All HPNP cleavage studies were carried out at 25℃and at PH 6.5-8.9 in 50mM buffer. UV-vis spectra and kinetic traces were recorded on a spectrophotometer equipped with a thermostated cell holder. The kinetic study was performed to measure the pseudofirst- order rate constants of the reactions by following the formation of p-nitrophenoxide. The pseudo-first-order rate constants were obtained from the slope of the absorbance versus time data. The pH dependence of the cleavage rate constants was assessed. The supramolecolar artifical nanoenzymes displayed significant rate enhancement of 105 times for the hydrolysis of HPNP.The hydrolytic kinetics of 2 -hydroxypropyl-4-nitrophenylp hosphate (HPNP)catalyzed by Cu(Ⅱ) and Zn(Ⅱ)complexes of supramolecolar artifical nanoenzyme and have been studied .The experiments demonstrated that this is an important model for the study of artifical metallic hydrolytic enzyme.