Introduction Molecular imprinting technology is a method tha

Introduction Molecular imprinting technology is a method that combines the material chemistry, biochemistry, and polymer chemistry and has the ability to specifically identify target analytes on shape, size and functional groups. Molecular imprinting polymer (MIP) is developed based on molecular imprinting technology, often referred as plastic antibody, have attracted much attention due to its unique facile preparation, high selectivity and sensitivity properties. Until now, it has been widely applied in many fields, such as solid-phase extraction, artificial adenosine receptor agonist and chemical sensors [1], [2]. MIP-based electrochemical sensors have the advantages from MIP and electrochemical sensor, including simple and low-cost instrument, easy signal quantification, rapidness, and so on, has been well applied in the detection of various substances such as metal ions, small molecules and proteins [3], [4], [5], [6], [7]. Usually, typical MIP include template molecules, functional monomers, crosslinking reagents, initiator and eluent. Among them, template molecules are target compounds in analytical processes, there are many types of molecularly imprinted polymer templates, such as ions, molecules, proteins, etc. Often an ideal template molecule should exhibit excellent chemical stability during the polymerization reaction, contain functional groups well adapted to assemble with functional monomers, and should not contain groups involved in or preventing polymerization [2]. So far, molecular imprinting method have been successfully applied for the recognition and detection of a wide variety of proteins, such as HIV-1 p24 [8], cTnІ [9], [10], PSA [11], [12] and so on. However usually, only those conformational stable proteins can be chosen as imprinted molecules, and a large number of proteins with unstable conformations such as gp120 have not been reported until now [13]. This may due to the target molecular can get into the hole during rebinding process only if they are in the correct posture as the imprinted state. For these proteins, the conformation in the resorbed solution is different from that in the polymerization, which will result in a bad blot. If the instability of the template can be solved, these proteins could be detected by molecularly imprinted electrochemical sensor. Gp120, the human immunodeficiency virus type 1 (HIV-1) envelope glycoprotein, is an important diagnostic marker of HIV-1 virus. The binding of HIV-1 gp120 to the human host cell surface receptor CD4 has been suggested as an important step in HIV infection and as a potential target for developing anti-HIV-1 therapy [14], [15], [16], [17]. Therefore, the detection of gp120 is very meaningful for the screening and prevention of HIV-1, develop a new method for fast and convenient monitoring of gp120 is highly desirable. However, the structure of gp120 is unstable, which leads to its inability to be detected by simple electrochemical methods. It can only be detected by some complicated methods, such as enzyme-linked immunosorbent assay [18]. During the research process, we found many studies have shown that the introduction of some molecule inhibitors can make the conformation of structurally variable protein into limited subset of states [19], [20]. NBD-556 is a kind of N-phenyl-Npiperidin-4-yl-oxalamide analogs, which can significantly inhibited syncytium formation. It is reported to block the interaction between the HIV-1 envelope glycoprotein gp120 and its receptor, CD4 [21], [22], [23]. When not bound, gp120 is in a relatively disordered state and different conformations can move between each other. The binding of NBD-556 can induce a large conformational change in the inner and outer regions of gp120, which make the conformational tend to be stable [21], [22]. Thus, we envisaged that small molecule inhibitors and structurally labile proteins could be used together as a blotting template to achieve the purpose of detecting labile proteins.
Experimental