Influence of induced fit in the interaction of Bacillus subtilis trp RNA-binding attenuator protein and its RNA antiterminator target oligomer.
In the presence of excess tryptophan, tryptophan-activated TRAP (trp RNA-binding attenuator protein) binds to a specific target in the trp-leader transcript, which induces the formation of a transcription terminator and transcription halts in the leader region. In the absence of tryptophan, TRAP does not bind RNA, an antiterminator forms, and the operon is expressed. Although the ternary complex involving TRAP (Bacillus stearothermophilus), tryptophan, and the RNA target has recently been crystallized, efforts to obtain structural data for the apo-form of TRAP (in any species) have not been successful. We have used multidimensional/multinuclear nuclear magnetic resonance (NMR) spectroscopy to probe the structure-function relationship in the TRAP-activated system, and have obtained high-resolution multidimensional/multinuclear NMR spectra of TRAP in all three of its functional states: tryptophan-free or apo-TRAP, tryptophan-activated TRAP, and tryptophan-activated TRAP-RNA ternary complex. Chemical shift perturbation analysis of the NMR data clarifies the interpretation of results obtained from previous crystal studies. Results presented herein demonstrate that tryptophan binding induces an essential structural change in TRAP that supports high-affinity binding of the RNA target oligonucleotide.
Flynn
Peter F
PF
Johnson Research Foundation and Department of Biochemistry and Biophysics, University of Pennsylvania, Philadelphia, Pennsylvania 19104, USA. pfflynn@mail.med.upenn.edu
Wendt
Alice
A
Gollnick
Paul
P
D020121:5' Untranslated Regions / Q000235:genetics / Q000378:metabolism; D001059:Apoproteins / Q000737:chemistry / Q000378:metabolism; D001412:Bacillus subtilis / Q000737:chemistry* / Q000235:genetics*; D001426:Bacterial Proteins / Q000737:chemistry / Q000378:metabolism; D001665:Binding Sites; D015964:Gene Expression Regulation, Bacterial; D009682:Magnetic Resonance Spectroscopy; D009841:Oligonucleotides / Q000235:genetics / Q000378:metabolism; D009876:Operon / Q000235:genetics; D011485:Protein Binding; D011487:Protein Conformation; D012329:RNA, Bacterial / Q000235:genetics* / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000737:chemistry* / Q000378:metabolism*; D013696:Temperature; D013728:Terminator Regions, Genetic / Q000235:genetics*; D014364:Tryptophan / Q000235:genetics*
D020121:5' Untranslated Regions; D001059:Apoproteins; D001426:Bacterial Proteins; D009841:Oligonucleotides; D012329:RNA, Bacterial; D016601:RNA-Binding Proteins; D014364:Tryptophan