Home > Research > Publications & Outputs > Monitoring Guanidinium-Induced Structural Chang...

Associated organisational unit

Text available via DOI:

View graph of relations

Monitoring Guanidinium-Induced Structural Changes in Ribonuclease Proteins Using Raman Spectroscopy and 2D Correlation Analysis

Research output: Contribution to Journal/MagazineJournal articlepeer-review

Published

Standard

Monitoring Guanidinium-Induced Structural Changes in Ribonuclease Proteins Using Raman Spectroscopy and 2D Correlation Analysis. / Brewster, Victoria L.; Ashton, Lorna; Goodacre, Royston.
In: Analytical Chemistry, Vol. 85, No. 7, 02.04.2013, p. 3570-3575.

Research output: Contribution to Journal/MagazineJournal articlepeer-review

Harvard

APA

Vancouver

Brewster VL, Ashton L, Goodacre R. Monitoring Guanidinium-Induced Structural Changes in Ribonuclease Proteins Using Raman Spectroscopy and 2D Correlation Analysis. Analytical Chemistry. 2013 Apr 2;85(7):3570-3575. doi: 10.1021/ac303265q

Author

Brewster, Victoria L. ; Ashton, Lorna ; Goodacre, Royston. / Monitoring Guanidinium-Induced Structural Changes in Ribonuclease Proteins Using Raman Spectroscopy and 2D Correlation Analysis. In: Analytical Chemistry. 2013 ; Vol. 85, No. 7. pp. 3570-3575.

Bibtex

@article{850594185cef4cec94c3ad88d0e652d6,
title = "Monitoring Guanidinium-Induced Structural Changes in Ribonuclease Proteins Using Raman Spectroscopy and 2D Correlation Analysis",
abstract = "Assessing the stability of proteins by comparing their unfolding profiles is a very important characterization and quality control step for any biopharmaceutical, and this is usually measured by fluorescence spectroscopy. In this paper we propose Raman spectroscopy as a rapid, noninvasive alternative analytical method and we shall show this has enhanced sensitivity and can therefore reveal very subtle protein conformational changes that are not observed with fluorescence measurements. Raman spectroscopy is a powerful nondestructive method that has a strong history of applications in protein characterization. In this work we describe how Raman microscopy can be used as a fast and reliable method of tracking protein unfolding in the presence of a chemical denaturant. We have compared Raman spectroscopic data to the equivalent samples analyzed using fluorescence spectroscopy in order to validate the Raman approach. Calculations from both Raman and fluorescence unfolding curves of [D](50) values and Gibbs free energy correlate well with each other and more importantly agree with the values found in the literature for these proteins. In addition, 2D correlation analysis has been performed on both Raman and fluorescence data sets in order to allow further comparisons of the unfolding behavior indicated by each method. As many biopharmaceuticals are glycosylated in order to be functional, we compare the unfolding profiles of a protein (RNase A) and a glycoprotein (RNase B) as measured by Raman spectroscopy and discuss the implications that glycosylation has on the stability of the protein.",
keywords = "ALPHA-CHYMOTRYPSIN, DENATURATION, UREA, PHARMACEUTICALS, GLYCOSYLATION, HYDROCHLORIDE, STABILITY, LYSOZYME",
author = "Brewster, {Victoria L.} and Lorna Ashton and Royston Goodacre",
year = "2013",
month = apr,
day = "2",
doi = "10.1021/ac303265q",
language = "English",
volume = "85",
pages = "3570--3575",
journal = "Analytical Chemistry",
issn = "0003-2700",
publisher = "American Chemical Society",
number = "7",

}

RIS

TY - JOUR

T1 - Monitoring Guanidinium-Induced Structural Changes in Ribonuclease Proteins Using Raman Spectroscopy and 2D Correlation Analysis

AU - Brewster, Victoria L.

AU - Ashton, Lorna

AU - Goodacre, Royston

PY - 2013/4/2

Y1 - 2013/4/2

N2 - Assessing the stability of proteins by comparing their unfolding profiles is a very important characterization and quality control step for any biopharmaceutical, and this is usually measured by fluorescence spectroscopy. In this paper we propose Raman spectroscopy as a rapid, noninvasive alternative analytical method and we shall show this has enhanced sensitivity and can therefore reveal very subtle protein conformational changes that are not observed with fluorescence measurements. Raman spectroscopy is a powerful nondestructive method that has a strong history of applications in protein characterization. In this work we describe how Raman microscopy can be used as a fast and reliable method of tracking protein unfolding in the presence of a chemical denaturant. We have compared Raman spectroscopic data to the equivalent samples analyzed using fluorescence spectroscopy in order to validate the Raman approach. Calculations from both Raman and fluorescence unfolding curves of [D](50) values and Gibbs free energy correlate well with each other and more importantly agree with the values found in the literature for these proteins. In addition, 2D correlation analysis has been performed on both Raman and fluorescence data sets in order to allow further comparisons of the unfolding behavior indicated by each method. As many biopharmaceuticals are glycosylated in order to be functional, we compare the unfolding profiles of a protein (RNase A) and a glycoprotein (RNase B) as measured by Raman spectroscopy and discuss the implications that glycosylation has on the stability of the protein.

AB - Assessing the stability of proteins by comparing their unfolding profiles is a very important characterization and quality control step for any biopharmaceutical, and this is usually measured by fluorescence spectroscopy. In this paper we propose Raman spectroscopy as a rapid, noninvasive alternative analytical method and we shall show this has enhanced sensitivity and can therefore reveal very subtle protein conformational changes that are not observed with fluorescence measurements. Raman spectroscopy is a powerful nondestructive method that has a strong history of applications in protein characterization. In this work we describe how Raman microscopy can be used as a fast and reliable method of tracking protein unfolding in the presence of a chemical denaturant. We have compared Raman spectroscopic data to the equivalent samples analyzed using fluorescence spectroscopy in order to validate the Raman approach. Calculations from both Raman and fluorescence unfolding curves of [D](50) values and Gibbs free energy correlate well with each other and more importantly agree with the values found in the literature for these proteins. In addition, 2D correlation analysis has been performed on both Raman and fluorescence data sets in order to allow further comparisons of the unfolding behavior indicated by each method. As many biopharmaceuticals are glycosylated in order to be functional, we compare the unfolding profiles of a protein (RNase A) and a glycoprotein (RNase B) as measured by Raman spectroscopy and discuss the implications that glycosylation has on the stability of the protein.

KW - ALPHA-CHYMOTRYPSIN

KW - DENATURATION

KW - UREA

KW - PHARMACEUTICALS

KW - GLYCOSYLATION

KW - HYDROCHLORIDE

KW - STABILITY

KW - LYSOZYME

U2 - 10.1021/ac303265q

DO - 10.1021/ac303265q

M3 - Journal article

VL - 85

SP - 3570

EP - 3575

JO - Analytical Chemistry

JF - Analytical Chemistry

SN - 0003-2700

IS - 7

ER -