Molecularly Imprinted Protein Recognition Cavities Bearing Exchangeable Binding Sites for Postimprinting Site-Directed Introduction of Reporter Molecules for Readout of Binding Events

Protein-imprinted cavities bearing exchangeable domains to be used for postimprinting fluorophore introduction to transform binding events into fluorescence changes were constructed in molecularly imprinted polymer (MIPs) matrixes prepared on glass substrates. Copolymerization was performed with acrylamide, <i>N</i>,<i>N</i>′-methylenebisaclylamide, and a newly designed functional group-exchangeable monomer, ({[2-(2-methacrylamido)­ethyldithio]­ethylcarbamoyl}­methoxy)­acetic acid (MDTA), in the presence of a model basic protein, lysozyme (Lyso); MDTA can interact with Lyso and assemble close to Lyso in the resulting polymer. After removal of Lyso, followed by a disulfide reduction to cleave the (ethylcarbamoylmethoxy)­acetic acid moiety from the MDTA residues, the exposed thiol groups within the imprinted cavities were modified by aminoethylpyridyldisulfide to be transformed into aminoethyl groups that function as active sites for amine-reactive fluorophores. Fluorescein isothiocyanate (FITC) was then coupled with the aminoethyl groups, yielding site specifically FITC-modified signaling imprinted cavities for Lyso binding. Because the in-cavity fluorescent labeling was achieved via a disulfide linkage, it was easy to remove, exchange, and/or replace amine-reactive fluorophores. This facilitated the screening of fluorophores to select the highest readout for binding events, replace fluorophores when photobleaching occurred, and introduce other functions. The proposed molecular imprinting process, combined with postimprinting modifications, is expected to provide an affordable route to develop multifunctional MIPs for specific detection of protein binding events

Supplementary material from Oriented, Molecularly Imprinted Cavities with Dual Binding Sites for Highly Sensitive and Selective Recognition of Cortisol

Materials, Apparatus, Organic synthesis, Preparation and characterization of the MIPs and the reference polymers, Possible sizes of β-CD complexes with FITC-BPA and TM1, Binding behaviors of FITC-BPA to MIPs, and Binding isotherms of cortisol in fluorescence-based competitive binding assay using the MIPs and the reference polymers for the estimation of apparent binding constants

Regioselective Molecularly Imprinted Reaction Field for [4 + 4] Photocyclodimerization of 2‑Anthracenecarboxylic Acid

Molecularly imprinted cavities have functioned as a regioselective reaction field for the [4 + 4] photocyclodimerization of 2-anthracenecarboxylic acid (2-AC). Molecularly imprinted polymers were prepared by precipitation polymerization of <i>N</i>-methacryloyl-4-aminobenzamidine as a functional monomer to form a complex with template 2-AC and ethylene glycol dimethacrylate as a crosslinking monomer. The 2-AC-imprinted cavities thus constructed preferentially bound 2-AC with an affinity greater than that toward structurally related 9-anthracenecarboxylic acid, 2-aminoanthracene, and unsubstituted anthracene. Moreover, from the four possible regioisomeric cyclodimers, they mediated the [4 + 4] photocyclodimerization of 2-AC specifically to the <i>anti</i>-head-to-tail (<i>anti</i>-HT) isomer. This indicates that the imprinted cavities accommodate two 2-AC molecules in an <i>anti</i>-HT manner, thereby facilitating the subsequent regioselective photocyclodimerization