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Extended Application of Gel-Permeation Chromatography by Spin Column

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Title Extended Application of Gel-Permeation Chromatography by
Spin Column
 
Creator Nath, Sangeeta
Brahma, Amrita
Bhattacharyya, Debasish
 
Subject Structural Biology & Bioinformatics
 
Description Separation of small volumes of proteins from unbound ligands or reequilibration with buffer by passing through a 1-ml Sephadex
G-50 column under mild centrifugal force is a popular technique. Here it has been demonstrated that other Sephadex matrix could
similarly be used for complete or partial separation of protein molecules. Proteins to be eluted at void volume are recovered near
quantitatively, while others are partly or almost completely retained depending on molecular size. Calibration curves using standard
proteins of Mw 12.5 to 440 kDa with Sephadex G-50–G-200 representing recovery versus molecular weight show profiles as expected
from the fractionation ranges of the column matrix. The procedure may be applied to follow protein association–dissociation reactions
if the molecular weights of the species concerned are known and a proper matrix exists for separating them. Equilibrium
unfolding transitions constructed with model proteins in presence of 0–8M urea using recovery as an index correspond to profiles
obtained from other physical measurements. This may be a convenient approach to follow change of protein hydrodynamic volume
quickly when a parallel methodology is not readily available.
 
Date 2003
 
Type Article
PeerReviewed
 
Format application/pdf
 
Identifier http://www.eprints.iicb.res.in/1194/1/35_AB_2003.pdf
Nath, Sangeeta and Brahma, Amrita and Bhattacharyya, Debasish (2003) Extended Application of Gel-Permeation Chromatography by Spin Column. Analytical Biochemistry, 320 (2). pp. 199-206.
 
Relation http://dx.doi.org/10.1016/S0003-2697(03)00302-6
http://www.eprints.iicb.res.in/1194/