Book Chapter

Absolute Quantitation of In Vitro Expressed Plant Membrane Proteins by Targeted Proteomics (MRM) for the Determination of Kinetic Parameters.

Carsten Rautengarten, Berit Ebert, Joshua L Heazlewood

Plant Membrane Proteomics: Methods and Protocols | Methods in Molecular Biology | Humana Press | Published : 2018

Abstract

The purification of a functional soluble protein from biological or in vitro expression systems can be problematic and the enrichment of a functional membrane protein for biochemical analyses can be a serious technical challenge. Recently we have been characterizing plant endomembrane nucleotide sugar transporters using a yeast expression system. However, rather than enriching these in vitro expressed proteins to homogeneity, we have been conducting biochemical characterization of these transport proteins in yeast microsomal fractions. While this approach has enabled us to estimate a variety of kinetic parameters, the accurate determination of the turnover number of an enzyme-substrate compl..

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