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.
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http://dx.doi.org/10.1039/b208487b | DOI Listing |
Biomol NMR Assign
January 2025
Department of Molecular Biology and Biophysics, University of Connecticut Health Center, Farmington, CT, 06030, USA.
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Escuela Superior de Física y Matemáticas, IPN S/N, Edificio 9 de la Unidad Profesional "Adolfo López Mateos", Col. Lindavista, Alc. Gustavo A. Madero, 07738, Mexico City, Mexico.
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Department of Food Science and Technology, Faculty of Agriculture, Ferdowsi University of Mashhad, Azadi Sq., Mashhad, Khorasan Razavi P.O. Box 9177948944, Iran. Electronic address:
Protein fibrillation complex mechanisms led to an emerging trend in research for years. The mechanisms behind whey protein isolate (WPI) fibrillation driven by divalent cations remained still a matter of speculation. All cations (Ca, Fe, Mg, and Zn) enhanced the microenvironment polarity through π-π stacking, and the amide I and II shifts confirmed the fibrillation.
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College of Food Science and Engineering, Northwest A&F University, Yangling, PR China. Electronic address:
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