Combination of noncovalent mass spectrometry and traveling wave ion mobility spectrometry reveals sugar-induced conformational changes of central glycolytic genes repressor/DNA complex.
The central glycolytic genes repressor (CggR) is a 37 kDa transcriptional repressor protein which plays a key role in Bacillus subtilis glycolysis by regulating the transcription of the gapA operon. Fructose-1,6-bisphosphate (FBP), identified as the effector sugar, has been shown to abolish the binding cooperativity of CggR to its DNA target and to modify the conformational dynamics of the CggR/DNA complex. In the present study, noncovalent mass spectrometry (MS) was used to obtain deeper insights into FBP-dependent CggR/DNA interactions. The effect of FBP binding on CggR alone and on CggR/DNA complexes was examined using automated chip-based nanoelectrospray MS and traveling wave ion mobility mass spectrometry (IM-MS). Our results revealed that tetrameric CggR dissociates into dimers upon FBP binding. Moreover, FBP binding to CggR/DNA complexes triggers disruption of intermolecular protein/protein interactions within the complex, significantly modifying its conformation as evidenced by a 5% increase of its collision cross section. For the first time, the use of IM-MS is reported to probe ligand-induced conformational modifications of a protein/DNA complex with an emphasis on the comparison with solution-based techniques.
Atmanene
Cédric
C
Laboratoire de Spectrométrie de Masse BioOrganique, Université de Strasbourg, IPHC, 25 rue Becquerel, 67087 STRASBOURG, France.
Chaix
Denix
D
Bessin
Yannick
Y
Declerck
Nathalie
N
Van Dorsselaer
Alain
A
Sanglier-Cianferani
Sarah
S
D001412:Bacillus subtilis / Q000201:enzymology / Q000235:genetics*; D001426:Bacterial Proteins / Q000737:chemistry*; D004268:DNA-Binding Proteins / Q000187:drug effects; D005635:Fructosediphosphates / Q000494:pharmacology*; D015964:Gene Expression Regulation, Bacterial; D006019:Glycolysis; D013058:Mass Spectrometry; D011487:Protein Conformation / Q000187:drug effects; D012097:Repressor Proteins / Q000737:chemistry*; D021241:Spectrometry, Mass, Electrospray Ionization
D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; D005635:Fructosediphosphates; D012097:Repressor Proteins; C029063:fructose-1,6-diphosphate