PMID:14962387

Citation	Ilag, LL, Westblade, LF, Deshayes, C, Kolb, A, Busby, SJ and Robinson, CV (2004) Mass spectrometry of Escherichia coli RNA polymerase: interactions of the core enzyme with sigma70 and Rsd protein. Structure 12:269-75
Abstract	The E. coli RNA polymerase core enzyme is a multisubunit complex of 388,981 Da. To initiate transcription at promoters, the core enzyme associates with a sigma subunit to form holo RNA polymerase. Here we have used nanoflow electrospray mass spectrometry, coupled with tandem mass spectrometry, to probe the interaction of the RNA polymerase core enzyme with the most abundant sigma factor, sigma70. The results show remarkably well-resolved spectra for both the core and holo RNA polymerases. The regulator of sigma70, Rsd protein, has previously been identified as a protein that binds to free sigma70. We show that Rsd also interacts with core enzyme. In addition, by adding increasing amounts of Rsd, we show that sigma70 is displaced from holo RNA polymerase, resulting in complexes of Rsd with core and sigma70. The results argue for a model in which Rsd not only sequesters sigma70, but is also an effector of core RNA polymerase.
Links	PubMed Online version:10.1016/j.str.2004.01.007
Keywords	DNA-Directed RNA Polymerases/metabolism; Electrophoresis, Polyacrylamide Gel; Escherichia coli/enzymology; Escherichia coli Proteins/metabolism; Repressor Proteins/metabolism; Sigma Factor/metabolism; Spectrometry, Mass, Electrospray Ionization
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