Peer-Reviewed Journal Details
Mandatory Fields
Groza, RC; Li, BY; Ryder, AG;
2015
July
Analytica chimica acta
Anisotropy resolved multidimensional emission spectroscopy (ARMES): A new tool for protein analysis
Published
Altmetric: 1WOS: 7 ()
Optional Fields
Fluorescence Anisotropy Multidimensional Protein Unfolding Chemometrics HUMAN-SERUM-ALBUMIN CELL-CULTURE MEDIA MULTIVARIATE CURVE RESOLUTION PRINCIPAL COMPONENT ANALYSIS FLUORESCENCE SPECTROSCOPY NOISE PERTURBATION CRYSTAL-STRUCTURE PRACTICAL ASPECTS EEM SPECTROSCOPY LOW-TEMPERATURES
886
133
142
Structural analysis of proteins using the emission of intrinsic fluorophores is complicated by spectral overlap. Anisotropy resolved multidimensional emission spectroscopy (ARMES) overcame the overlap problem by the use of anisotropy, with chemometric analysis, to better resolve emission from different fluorophores. Total synchronous fluorescence scan (TSFS) provided information about all the fluorophores that contributed to emission while anisotropy provided information about the environment of each fluorophore. Here the utility of ARMES was demonstrated via study of the chemical and thermal denaturation of human serum albumin (HSA).Multivariate curve resolution (MCR) analysis of the constituent polarized emission ARMES data resolved contributions from four emitters: fluorescence from tryptophan (Trp), solvent exposed tyrosine (Tyr), Tyr in a hydrophobic environment, and room temperature phosphorescence (RTP) from Trp. The MCR scores, anisotropy, and literature validated these assignments and showed all the expected transitions during HSA unfolding. This new methodology for comprehensive intrinsic fluorescence analysis of proteins is applicable to any protein containing multiple fluorophores. (C) 2015 Elsevier B.V. All rights reserved.
http://www.sciencedirect.com/science/article/pii/S0003267015008065
10.1016/j.aca.2015.06.011
Grant Details
RCG was supported by an ‘EMBARK Initiative’ Postgraduate Scholarship from the Irish Research Council. We also thank Jeffrey Comerford and Ursula Tems of Agilent Technologies (Mulgrave Victoria, Australia) for the loan of a fluorescence spectrometer.
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