Finding reliable phenotypes and detecting artefacts among in vivo and in vitro assays to characterize the refractory transcriptional activator Sxy (TfoX) in Escherichia coli

Abstract

14The Sxy (TfoX) protein is required for expression of a distinct subset of the genes regulated 15 by the cAMP receptor protein (CRP) protein purification protocols and quality control steps for nickel affinity column purification, 31 protein mass spectrometry revealed the enrichment of additional DNA-binding proteins in nickel 32 column eluates, presenting a probable source of artefactual protein-protein and protein-DNA 33 interaction results. These findings highlight the importance of extensive controls and phenotypic 34 assays for the study of poorly characterized and recalcitrant proteins like Sxy.