Abstract:
:The resolution of complex protein mixtures by discontinuous buffer SDS-PAGE is accomplished by their concentration into thin bands in the stacking gel, followed by their separation during migration through the resolving gel. Recombinant human interferon-inducible protein-10 (IP-10), a 10-kDa C-X-C chemokine with four cysteines, aggregated during the stacking phase of SDS-PAGE and generated a band with an apparent molecular mass of 18 kDa. This aggregation depended on the presence of reduced sulfhydryl residues on IP-10, on the amount of loaded protein, and on the concentration of the ammonium persulfate used to polymerize the stacking gel. The aggregation of IP-10 could be prevented by reduction of its sulfhydryls with dithiothreitol followed by irreversible blockade with iodoacetamide. These methods may be useful in the prevention of aggregation of sulfhydryl-containing proteins during SDS-PAGE, especially when large quantities are analyzed to assess their purity.
journal_name
Biotechniquesjournal_title
BioTechniquesauthors
Crow MK,Karasavvas N,Sarris AHdoi
10.2144/01302st04subject
Has Abstractpub_date
2001-02-01 00:00:00pages
311-6issue
2eissn
0736-6205issn
1940-9818journal_volume
30pub_type
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