title: The characterization of biomolecular secondary structures by surface plasmon resonance.
abstract: Recently there has been considerable interest in using surface plasmon resonance (SPR) for the measurement of conformational changes of immobilized biomolecules that are induced by an exogenous analyte. While a number of studies have shown the analytical utility of such measurements, there has been no report which characterizes the specific secondary structure that actuates the change in SPR signal. The use of SPR to indicate the type of secondary structure present in two immobilized polypeptides, poly-L-lysine (PL) and poly-L-glutamic acid (PGA), and a globular protein, concanavalin A (Con A) is described in this report. The PL, PGA and Con A were modified with N-succinimidyl 3-(2-pyridyldithiol) propionate (SPDP) to introduce disulfide groups to facilitate the attachment onto gold-coated surfaces via self-assembly. Ethanol and 2,2,2-trifluoroethanol (TFE) were used to induce changes in the secondary structure of the immobilized polypeptides and the protein respectively. Using both circular dichroism (CD) and Fourier transform infrared (FTIR) spectroscopies, it has been demonstrated that it is possible to correlate the signal changes observed in SPR to the secondary conformation of the biomolecule. Both CD and FTIR showed that a decrease in SPR signal corresponded to a high content of beta, turn or unordered structures while an increase corresponded to a high alpha-helical content. The sensitivity of the SPR technique is comparable to that obtained in solution with CD and FTIR spectroscopies. These results are the first demonstration that SPR can be used to characterize secondary structures. There is potential, therefore, for SPR to be used as a technique to study secondary conformational changes of immobilized polypeptides and proteins.
authors.affiliation: School of Chemical Sciences and Pharmacy, University of East Anglia, Norwich, UK NR4 7TJ.
authors.forename: Lee May
authors.initials: LM
authors.lastname: May
authors.forename: David A
authors.initials: DA
authors.lastname: Russell
mesh_terms: D002942:Circular Dichroism; D003208:Concanavalin A / Q000737:chemistry; D011099:Polyglutamic Acid / Q000737:chemistry; D011107:Polylysine / Q000737:chemistry; D017433:Protein Structure, Secondary; D017550:Spectroscopy, Fourier Transform Infrared; D020349:Surface Plasmon Resonance
chemical_list: D003208:Concanavalin A; D011107:Polylysine; D011099:Polyglutamic Acid