Proteome purification

In the March 5 Proceedings of the National Academy of Sciences, Pascal Braun and colleagues describe a methodology to perform high-throughput purification of human proteins using several affinity-tags (Proc Natl Acad Sci USA 2002, 99:2654-2659).Braun et al. exploited the Gateway recombinational cloning system that employs a versatile 'master' vector for easy transfer into different expression vectors. They chose a test set of 32 human genes and expressed them in four different vectors with diffe

Jonathan Weitzman(jonathanweitzman@hotmail.com)
Mar 7, 2002

In the March 5 Proceedings of the National Academy of Sciences, Pascal Braun and colleagues describe a methodology to perform high-throughput purification of human proteins using several affinity-tags (Proc Natl Acad Sci USA 2002, 99:2654-2659).

Braun et al. exploited the Gateway recombinational cloning system that employs a versatile 'master' vector for easy transfer into different expression vectors. They chose a test set of 32 human genes and expressed them in four different vectors with different affinity tagsL: the His6tag, calmodulin-binding peptide, glutathione-S-transferase or maltose-binding protein. Using different denaturing and nondenaturing purification conditions they were able to isolate around 80% of expressed proteins.

They demonstrated that the system can be easily scaled-up, and then applied the same approach to a set of 336 human cDNAs with a success rate of around 60%. This approach allows for optimization of protein purification on a proteome-scale.

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