The methylotrophic yeast Pichia pastoris is now an established expression system for the production of recombinant protein for nuclear magnetic resonance (NMR) studies. It is capable of expressing high levels of heterologous proteins and possesses the ability to perform many of the posttranslational modifications of higher eukaryotes. Here, we describe efficient methods for the production of uniformly 13C,15N-labeled proteins in shake flasks and of uniformly 13C,15N-labeled and 2H,13C,15N-labeled proteins in fermenters. We also provide details of two chromatographic procedures, cation exchange and concanavalin A lectin affinity, that have proved useful in purifying P. pastoris-expressed proteins for NMR studies.
|Title of host publication||Protein NMR techniques|
|Place of Publication||Totowa|
|Number of pages||17|
|Publication status||Published - 2004|
|Name||Methods in molecular biology|
|Publisher||Humana Press Inc|