Function of two GSA isoforms in Arabidopsis thaliana

As proof of interdependent interaction between GSAT, GluTR, and GBP, a crosslinking mass spectrometry experiment was conducted with bilateral and trilateral combinations of the three respective recombinant proteins. Apart from the known GBP – GluTR interaction, combined assays for mutual interaction of GSAT either with GluTR or GBP were carried out and ultimately a crosslinking experiment was performed including all three proteins. The proteins of these assays were subsequently cleaved by trypsin and analysed by mass spectrometry to identify the crosslinked peptides of the three proteins in all possible of dual and triple combinations. This approach enabled the detection of the interrelation between all three proteins.