Screening of novel peptides that specifically interact with vitamin D bound biocomplex proteins

The majority of the vitamin D that is present in the blood binds to vitamin D binding protein (VDBP) and circulates in the form of a complex (VDBP-Complex). Knowing the level of vitamin D in the body is crucial for vitamin D-related treatments so that the right dosage of vitamin D can be given. In other words, it is essential to distinguish between the protein VDBP and the complex form bound to vitamin D. As a novel way for the detection of VDBP-Complex, a more effective phage display methodology was applied in this study along with the addition of two approaches. In order to screen a sequence specific to the target only, the pre-binding method and after-binding method were performed. VDBP-Complex was directly coated on the petri dishes. In order to select phages that specifically bind to the VDBP-Complex, random phages were attached, and selected by 7 times of biopanning. Individual DNA sequences were analyzed for each biopanning to find specific peptide sequences for VDBP-Complex. The affinity of binding phages was verified by ELISA assay using an anti-M13 antibody. The phage having a sequence of SFTKTSTFTWRD (called as M3) has shown the highest binding affinity to VDBP-Complex. As a result of the removal test of VDBP-Complex using magnetic beads conjugated with M3 peptide, it was confirmed that significant decrease of VDBP-Complex. The unique characteristic of the M3 sequence was confirmed through a sequence-modified peptide (SFT motif). That is, it is expected that the M3 peptide may be used to determine the vitamin D levels in the blood.

Automated summary

This study applied a novel method for the detection of VDBP-Complex in the blood, and identified a phage with the highest binding affinity to VDBP-Complex, which has a sequence of SFTKTSTFTWRD (called M3). The significant decrease of VDBP-Complex was confirmed through the removal test using magnetic beads conjugated with M3 peptide. It is expected that the M3 peptide may be used to determine the vitamin D levels in the blood.