Immobilization of RNase S-Peptide on a single-stranded DNA-fixed gold surface and effective masking of its surface by an acridinyl poly(ethylene glycol)

Abstract

Oligonucleotide-peptide conjugate was synthesized by coupling of RNase S-peptide to a 24-mer single-stranded DNA (ssDNA) oligonucleotide to be immobilized on its complementary ssDNA oligonucleotide-fixed gold surface of sensor chip or electrode. Immobilization of on the ssDNA-fixed gold surface through DNA duplex formation was confirmed by quartz crystal microbalance (QCM) and electrochemical measurements. After treating with a synthetic acridinyl poly(ethylene glycol) (APEG), specific interaction of S-protein with the S-peptide immobilized on the gold surface was demonstrated by QCM without nonspecific adsorption of unrelated proteins such as BSA and RNase A at the surfaces. This result suggested that the acridine parts of APEG could bind to the DNA duplex on the gold surface and the poly(ethylene glycol) parts were fastened on the surface to resist the adsorption of proteins. Thus, the combination of oligonucleotide-peptide conjugate, ssDNA-fixed chip and APEG with effective masking property provides a new tool for the analysis of specific peptide-protein interactions without disturbance by other unrelated proteins.

Biomolecular interactions and tools for their recognition: focus on the quartz crystal microbalance and its diverse surface chemistries and applications

“…One QCM, for example, uses an immobilized peptide to detect the presence of a specific protein in solution. 91 More specifically, the electrode surface is functionalized with a mercaptohexanol SAM to attach ssDNA, to which complementary, RNase S-peptide-conjugated ssDNA binds. A final layer of acridinyl polyethylene glycol (APEG) is used to mask the gold electrode to prevent nonspecific adsorption of proteins onto the electrode surface (via the PEG portion of APEG) and to bind dsDNA (via the acridine moiety).…”

“…69 In contrast, one QCM chip directly detects adsorption of albumin and fibrinogen onto an electrode that is modified by an alkanethiol SAM, 86 while another thiol SAM-based QCM uses immobilized S-peptide to detect S-protein. 91 A different QCM used an immobilized peptide probe, however this time the QCM chip was modified by an amine SAM and detected human interferon-b. 96 Alternatively, with an immobilized peptide A probe, an amide SAM-modified QCM detected profilin protein.…”

Identification of Trimeric Peptides That Bind Porcine Parvovirus from Mixtures Containing Human Blood Plasma

“…A spacer arm of two sets of two ethylene glycol units separated by a peptide bond (26) (designated AEEA‐AEEA by Peptides International) was added to the library to increase the chances of finding a peptide ligand that bound to a conserved area on the virus surface. This spacer arm separated the peptide approximately 15 Å from the undisclosed spacer on the Toyopearl resin.…”

Recent advances in electrochemical DNA hybridization sensors

“…Takenaka and co-workers employed a standard thiol linked dsDNA electrode and created a peptide analysis chip by 5 0 -modifiying the captured DNA target with RNase S-peptide. 82 DNA hybridization was indirectly electrochemically monitored using Fc modified naphthalene diimide. Hybrids were exposed to acridine modified polyethylene glycol (PEG) molecules that intercalated into the duplexes.…”