Functionalized receptors for efficient epitope recognition

Trinity University received $535,867 from the National Institute of General Medical Sciences (NIGMS) under the Biomedical Research and Research Training program (CFDA 93.859) for a three-year project grant (September 1, 2025 through August 31, 2028) focused on developing functionalized synthetic receptors for protein recognition. The research aims to create small monofunctional synthetic receptors derived from cucurbit[n]urils that can bind to minimal epitope tags (2-3 amino acid residues) with high affinity and specificity. These receptors will provide subnanomolar binding affinity while remaining functional under physiological conditions, including in serum environments.

The project deliverables include: (1) optimization of epitope sequences for enhanced recognition; (2) integration of these epitopes into recombinant proteins and human insulin; and (3) development of monofunctional receptor derivatives with multiple applications including affinity purification, protein sensing, and chemical labeling capabilities. The synthetic receptors will enable on-demand protein release through small molecule competitors while minimizing impact on the structure and activity of target proteins. This research builds upon established foundational work in sequence-selective peptide and protein recognition and is expected to establish core competencies applicable to existing and emerging biomedical applications.

Abstract

Biomedical science relies heavily on methods for attaching ligand binding sites to proteins. Such epitope tags are important for protein affinity purification, for improving protein solubility, and for adding functionality to proteins such as fluorescent labels or sites for chemical modification. The proposed research aims to develop synthetic receptors that target proteins engineered with efficient epitopes comprising only 2-3 amino acid residues. The epitopes are bound site-selectively and with subnanomolar affinity by small (1-2 KDA) monofunctional synthetic receptors with added ability to bind, fluoresce, or attach to solid supports. The receptors are stable and functional in physiological conditions, including serum, and proteins bound to these receptors can be released on demand upon the addition of small molecule competitors. The proposed aims will: 1) further optimize the epitopes; 2) install the epitopes into recombinant proteins and human insulin; and 3) develop monofunctional derivatives of the synthetic receptor to enhance binding and to enable affinity purification and protein sensing. The proposed epitopes will add function to target proteins while having minimal impact on structure and activity. The proposed work builds on the rules for sequence-selective recognition of peptides and proteins by cucurbit[n]urils established in the undergraduate research lab of the PI. Many research groups have developed applications built on these rules, and we anticipate that the proposed research will establish a core competency that should facilitate existing biomedical applications and inspire new ones.