A quantitative, high-throughput screen for protein stability.
In proteomic research, it is often necessary to screen a large number of polypeptides for the presence of stable structure. Described here is a technique (referred to as SUPREX, stability of unpurified proteins from rates of H/D exchange) for measuring the stability of proteins in a rapid, high-throughput fashion. The method uses hydrogen exchange to estimate the stability of microgram quantities of unpurified protein extracts by using matrix-assisted laser desorption/ionization MS. The stabilities of maltose binding protein and monomeric lambda repressor variants determined by SUPREX agree well with stability data obtained from conventional CD denaturation of purified protein. The method also can detect the change in stability caused by the binding of maltose to maltose binding protein. The results demonstrate the precision of the method over a wide range of stabilities.
Ghaemmaghami
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Department of Biochemistry, Duke University Medical Center, Durham, NC 27710, USA.
Fitzgerald
M C
MC
Oas
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TG
D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D010582:Bacteriophage lambda / Q000737:chemistry; D002352:Carrier Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins; D008320:Maltose / Q000235:genetics / Q000378:metabolism*; D058619:Maltose-Binding Proteins; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012097:Repressor Proteins / Q000737:chemistry* / Q000235:genetics; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*; D014764:Viral Proteins / Q000737:chemistry* / Q000235:genetics; D054334:Viral Regulatory and Accessory Proteins
D001426:Bacterial Proteins; D002352:Carrier Proteins; D004268:DNA-Binding Proteins; D058619:Maltose-Binding Proteins; D011993:Recombinant Fusion Proteins; D012097:Repressor Proteins; D014764:Viral Proteins; D054334:Viral Regulatory and Accessory Proteins; C016649:phage repressor proteins; D008320:Maltose