PAGE MW estimates

douglas l feinstein dlfeins at cumc.cornell.edu
Tue Mar 8 10:13:27 EST 1994


			If a protein contains a high content of hydrophobic residues, or
particularly strong hydrophobic domains (e.g. a membrane bound or
integral membrane protein), it may not unfold completely even in the
presence of 1% SDS. One can get an idea of hydrophobic nature by carrying
out PAGE In the presence of SDS and with varying concentration so f Urea
(from 0 to 8M). Urea should increase the likelihood of protein unfolding,
and many proteins will run faster in the presence of urea, closer totheir
true MW. Another factor may be that the protein does not bind equal
amounts of SDS per unit or protein, which can be due to unusual AA
sequence runs, and again hydrophobic nature. 
	Another way to better estimate MW by PAGE is to carry out Ferguson plot
analysis. Determine the mobility of the proteins, and markers, at
different concentations of acrylamide. SLopes of mobility versus the
standard give very accurate measures of MW. The intercepts of these
curves can be interpreted in terms of amount of SDS bound per unit
protein and/or the overall shape of the migrating moleucle (i.e. length
to width ratio of the  ideal cyllinder).

dlf
Doug Feinstein																	 |  Voice:   212 570-2900
Dept Neurobiology															|	Fax:      212 988-3672
Cornell University Medical College    |  E-mail: dlfeins at cumc.cornell.edu
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