TY  - JOUR
TI  - Expression and Purification of Arabidopsis Transmembrane Protein BCM1 in  Saccharomyces cerevisiae
AU  - Wang, Peng 
AU  - Grimm, Bernhard 
VL  - 10
IS  - 18
PY  - 2020
DA  - 2020/09/20
SP  - e3758
C1  - Bio-protocol 2020;10:e3758
DO  - 10.21769/BioProtoc.3758
UR  - https://doi.org/10.21769/BioProtoc.3758
AB  - Heterologous expression and purification of transmembrane proteins have remained a challenge for decades hampering detailed biochemical and structural characterization of key enzymes and their interacting regulators in multiple metabolic pathways. An in-depth study on the newly identified Arabidopsis thaliana integral membrane protein BALANCE OF CHLOROPHYLL METABOLISM 1 (BCM1) showed a stimulatory effect of the BCM1 on magnesium chelatase, the first enzyme of chlorophyll biosynthesis, through interaction with the GENOMES UNCOUPLED 4 (Wang et al., 2020). Here, we report a detailed and optimized method for heterologous expression and purification of His-tagged BCM1 in Saccharomyces cerevisiae. Following this method, we obtained native BCM1 used for in vitro enzymatic assay of magnesium chelatase (Wang et al., 2020). Currently, the crystallization studies of the BCM1 are underway. This protocol could be adapted to purify BCM1-like transmembrane proteins from eukaryotic organisms for enzymatic and structural studies.
KW  - A transmembrane protein
KW  - Expression and purification of BCM1
KW  - Saccharomyces cerevisiae
KW  - Arabidopsis thaliana
JF  - Bio-protocol
SN  - 2331-8325
PB  - Bio-protocol LLC.
BIO101  - False