Protein-ligand binding detected using ultrafiltration Raman difference spectroscopy.

Protein-ligand binding detected using ultrafiltration Raman difference spectroscopy. Research Abstract Details 

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Protein-ligand binding detected using ultrafiltration Raman difference spectroscopy. Abstract Text:

Yong Xie,Dongmao Zhang,Dor Ben-Amotz,Yong Xie,Dongmao Zhang,Dor Ben-Amotz,

A new ultra-filtration-Raman-difference (UFRD) method facilitates the tag-free screening and quantitation of protein-ligand binding constants. The method relies on drop-coating-deposition-Raman (DCDR) combined with ultrafiltration and difference spectroscopy. Ultrafiltration is used to remove free (unbound) ligands from pre-equilibrated protein/ligand solutions. Difference DCDR spectroscopy is used to detect binding-induced vibrational spectral changes obtained from proteins with and without a bound ligand. The capabilities of the UFRD method are demonstrated using the binding of 2,4-dinitrophenol (DNP) to transthyretin (TTR), as well as preliminary measurements in several other systems. The UFRD results clearly reveal DNP spectral features induced by binding to TTR and confirm that only a 1:1 complex is formed even under 10-fold excess DNP conditions. The UFRD method is shown to be most useful when applied to strongly Raman active ligands (such as aromatic compounds). Weakly Raman-active ligands (such as sugars) are typically not compatible with UFRD detection (unless they produce a sufficiently large binding-induced change in protein secondary structure). Theoretical predictions suggest that UFRD may be used to screen binding events with a dissociation constant cut-off of the order of 10muM, and perhaps also to quantify dissociation constants in the 100nM to 100muM range.

Protein-ligand binding detected using ultrafiltration Raman difference spectroscopy. Publishing Authors By Initials

Y Xie,D Zhang,D Ben-Amotz,Y Xie,D Zhang,D Ben-Amotz,

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Protein-ligand binding detected using ultrafiltration Raman difference spectroscopy. Journal Published:

PUBLICATION TYPE: Journal Article

Journal: Analytical biochemistry

VOLUME: 373

Page Numbers: 154-60

Journal Abbreviation: Anal. Biochem.

ISSN: 0003-2697

DAY: 4

MONTH: 11

YEAR: 2007

Protein-ligand binding detected using ultrafiltration Raman difference spectroscopy. Information

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LANGUAGE: eng

NlmUniqueID: 370535

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Grant and Affiliation Information for Protein-ligand binding detected using ultrafiltration Raman difference spectroscopy.

AFFILIATION: Purdue University, Department of Chemistry, West Lafayette, IN 47907, USA.

Country: United States

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MEDLINETA: Anal Biochem

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