Development of a peptide-based KIM-1 biosensor utilizing computationally designed CCT3-derived peptides

Abstract

The kidney injury molecule-1 (KIM-1) serves as a well-recognized biomarker for kidney injury, making the establishment of accurate and sensitive detection methods highly valuable for clinical applications. This research involved computational design and analysis of protein-binding peptide fusion; MBP-CCT3-BP03 and MBP-CCT3-BP04 were engineered to bind to KIM-1. Because peptides are generally small in size, both designed binding peptides were fused with maltose binding protein (MBP) to assist in their expression and biosensor fabrication. The affinity study and the development of biosensors were conducted electrochemically. The molecular docking binding affinity of CCT-BP03 and CCT3-BP04 against KIM-1 was predicted at −8.8 kJ/mol and − 8.1 kJ/mol respectively, while their calculated MM/PBSA were recorded at −45.70 kJ/mol and − 36.1 kJ/mol, respectively. In addition. The MD simulation structural analysis reveals that MBP-CCT3-BP04 exhibits better structural stability and folding during the 100 ns simulation while interacting with KIM-1, hence it was chosen for the development of the KIM-1 biosensor. Under optimal conditions, the developed biosensor showed a linear correlation of MBP-CCT3-BP04 related to the concentration of KIM-1 with a detection limit of 1.02 ng/mL, where it falls within the clinical range of KIM-1 in urine. The designed MBP-CCT3-BP04 also demonstrates high specificity and selectivity in both the spiked buffer and the simulated urine sample. Overall, this research establishes the viability of using computational procedure in designing binding peptides as well as employing protein-binding peptide fusion approach for biosensor development. This approach provides a practical framework for creating cost-effective and reliable KIM-1 detection platforms that could have significant clinical utility.