Designed Multifunctional Isopeptide for Enhanced Annexin A1 Biosensing Based on Peptide–Protein Interactions in Human Blood

Abstract

Specific peptide−protein interactions play an important role in biosensing systems based on functional peptides; however, the non-specific interactions with unrelated biomolecules and poor proteolytic stability restrict the clinical application of natural peptides. Here, we leveraged a self-designed multifunctional isopeptide (MISP) to construct an electrochemical biosensing platform for annexin A1 (ANXA1) detection in human blood. The MISP was designed to contain two parts: an antifouling cyclotide cyclo-C(EK) 4 and a Damino acid-containing carbohydrate-mimetic recognizing peptide IF-7 (D-IF7) connected by the isopeptide bond. We have discussed the properties of the cyclotide and illustrated its unique advantage over the natural linear antifouling peptides by molecular dynamics simulations, and the results were further confirmed by dissipative quartz crystal microbalance (QCM-D). In addition, through electrochemical experiments and fluorescence imaging experiments, we demonstrated that the MISP-based biosensor possessed excellent antifouling ability and proteinase hydrolysis stability. Interestingly, the assaying results of the MISP-biosensor were consistent with those of the commercial ANXA1 kits in a variety of healthy and ANXA1-upregulated clinical blood samples, and, more importantly, for the analysis of blood samples with lower ANXA1 expressions, the sensing capability of the biosensor was greatly superior to that of the kits because of the lower detection limit of the MISPbiosensor. This biosensing platform based on the designed MISP offers enormous potential for achieving accurate biomarker detection with robust operation in complex biological samples.

Phospholipid Bilayer Integrated with Multifunctional Peptide for Ultralow-Fouling Electrochemical Detection of HER2 in Human Serum

“…The images in Figure S3A exhibit the morphology of SA-doped PEDOT showing a microporous structure, which was consistent with our previous work. 24 The presence of DSPE-PEG was clearly observed after DSPE-PEG modification (Figure S3B). These results verified the successful construction of the biosensor.…”

“…First of all, PEDOT/GCE was obtained by electropolymerization at a constant potential of 1.1 V for 20 s in a solution containing 3.0 mg mL −1 SA and 9.0 mM EDOT monomer. 24 Subsequently, the carboxyl group of SA in the PEDOT/GCE was activated for 0.5 h by EDC/ NHS solution (containing 0.4 M EDC and 0.1 M NHS), and then 20 μL of DSPE-PEG solution (10.0 mg mL −1 ) was dropped on the PEDOT/GCE surface and covered with a wet beaker for 2 h, and the DSPE-PEG (with amino group) was then attached to the modified electrode (with carboxyl group) via amide bonding to obtain the DSPE-PEG/PEDOT/GCE. Afterward, the prepared DSPE-PEG/PEDOT/GCE was soaked in 2.0 mM sulfo-SMCC aqueous solution for 1 h at room temperature.…”

Designed Polyhydroxyproline Helical Peptide with Ultrarobust Antifouling Capability for Electrochemical Sensing in Diverse Complex Biological Fluids

“…The presence of proteases accountable for peptide hydrolysis can present a formidable obstacle to the durability and efficacy of peptide-based biosensors in biological fluids. The significance of enhancing the resistance to protease degradation of peptide biosensors was thus appropriately emphasized . Indeed, it is widely recognized that D-peptides possess excellent protease tolerance due to the fact that most proteases specifically recognize and hydrolyze peptides containing the natural l -amino acids .…”

“…The significance of enhancing the resistance to protease degradation of peptide biosensors was thus appropriately emphasized. 21 Indeed, it is widely recognized that D-peptides possess excellent protease tolerance due to the fact that most proteases specifically recognize and hydrolyze peptides containing the natural L-amino acids. 6 Hence, by constructing the PHHP using exclusively D-type amino acids, inherent protease tolerance can be achieved.…”

Antifouling Electrochemiluminescence Biosensor Based on Bovine Serum Albumin Hydrogel for the Accurate Detection of p53 Gene in Human Serum

“…Generally, water and other liquids exhibit smaller contact angles on hydrophilic surfaces, making it difficult for nonspecific proteins to adhere, thus reducing interference on biosensors. Hydrogels crosslinked with other materials typically exhibit high biocompatibility and conductivity, such as bovine serum albumin (BSA), polypeptide, , and polyaniline. , These materials have low toxicity and immunogenicity in practical samples, allowing contact without adverse reactions. Therefore, incorporating hydrogel coatings in electrochemical biosensors enhances biocompatibility and reduces potential biological toxicity risks.…”