The Fabrication Of Calcium-induced Nanochannels Based On Smart Polymers

In nature,with high selectivity to specific ions,ion channels play key roles in controlling ion transport between cells and their surroundings.Inspired by nature,scientists utilize solid substrates with nanochannels and special modification materials to simulate the performance of ion channels in cell membrane.Similar to biological nanochannels,artificial nanochannels can respond to subtle changes in the environment and have good designability and expandability,which have good application potential and development prospects in the fields of biosensing,drug controllable release,bio-separation.In the past few decades,some important achievements have been achieved in the field of conventional voltage-gated calcium ion channels.However,These modulation strategies mainly rely on the surface charge changes of the nanochannels to achieve“on-off”switching,which are much different from the ion channels in vivo whose gating behaviors are mainly achieved by the conformational transition of channel proteins,and makes the traditional voltage-gated calcium channel not the best choice for the current application and development of the biological field.Therefore,making full use of the responsiveness and functionality of intelligent polymers for the further development of new artificial nanochannels is particularly important for the current biological field.In this work,inspired by the Ryanodine Receptor(Ry R)ion channel,we used the conformational transformation of the polymer to simulate the conformational change of the channel protein during the transportation process of ions.Designed and prepared a new type of calcium responsive polymer.And porous anodic alumina(PAA)with adjustable membrane thickness and high porosity is selected as the substrate to construct the nanochannel system.Finally,a unique calcium ion regulated channel with bionic properties was obtained.The specific work includes the following aspects:To begin with,a calcium responsive peptide sequence(DDDEEKC)was selected.The binding affinity of the DDEEKC with Ca Cl₂ was evaluated by isothermal titration calorimetry(ITC)and ¹³C NMR titration experiments.Apart from that,Circular dichroism(CD)and molecular dynamic calculations was applied to investigate conformation changes of DDDEEKC under the induce of calcium,from?-helix to random coil.Based on the previous works of our team,a three–component copolymer was designed according to a“Recognition–Mediation–Main chain”concept,in which the hepta-peptide DDDEEKC worked as a Ca²⁺recognition unit,CF₃-PT functioned as a hydrogen bond mediation unit and PNI provided a flexible polymer main chain.The interaction abilities between monomers was verified by fluorescence titration.Then,the polymer was grafted onto the surface of silicon chips,QCM chips and gold electrodes to investigate the adsorption capacity of the polymer film to calcium ions as well as the globule-to-coil transition of the polymer chain.Finally,a calcium responsive artificial nanochannel system was constructed,and its properties were characterized by picoammeter.The results show that the nanochannel systerm could achieve intelligent response ability at an ultra-low concentration(10pmol).At low concentration,the polymer in the nanochannel is in a contracted state and the channel is open wide.As the concentration gradually increased,the polymer chains which could response to calcium gradually changed to a swollen one and the channel gradually closed,resulting in a significantly smaller transmembrane current.The highest change ratio of transmembrane current could achieve as high as 47.5%.In addition,the channel had good ion selectivity and maintained good regulation ability after seven rounds of cyclic test.This closely bionic design inspired from biological ion channels will enlighten more biomimetic nanochannels based on intelligent polymers,and its excellent performance can promote the development of important applications of tissue engineering field.