Measurement of internal movements within the 30 S ribosomal subunit using Förster resonance energy transfer.
We have used Förster resonance energy transfer (FRET) to study specific conformational changes in the Escherichia coli 30 S ribosomal subunit that occur upon association with the 50 S subunit. By measuring energy transfer between 13 different pairs of fluorescent probes attached to specific positions on 30 S subunit proteins, we have monitored changes in distance between different locations within the 30 S subunit in its free and 50 S-bound states. The measured distance changes provide restraints for modeling the movement that occurs within the 30 S subunit upon formation of the 70 S ribosome in solution. Treating the head, body, and platform domains of the 30 S subunit as simple rigid bodies, the lowest-energy solution converges on a model that satisfies each of the individual FRET restraints. In this model, the 30 S subunit head tilts towards the 50 S subunit, similar to the movement found in comparing 30 S subunits and 70 S ribosomes from X-ray and cryo-electron microscope structures, and the platform is predicted to undergo a clock-wise rotation upon association.
Hickerson
Robyn
R
Center for Molecular Biology of RNA and Department of Molecular, Cell and Developmental Biology, Sinsheimer Laboratories, University of California, Santa Cruz, CA 95064, USA.
Majumdar
Zigurts K
ZK
Baucom
Albion
A
Clegg
Robert M
RM
Noller
Harry F
HF
D001665:Binding Sites; D020285:Cryoelectron Microscopy; D018360:Crystallography, X-Ray; D004926:Escherichia coli / Q000378:metabolism*; D031541:Fluorescence Resonance Energy Transfer; D008958:Models, Molecular; D011487:Protein Conformation; D012269:Ribosomal Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D012270:Ribosomes / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
D012269:Ribosomal Proteins; C040469:ribosomal protein S30