Book of Abstracts: Albany 2007

category image Albany 2007
Conversation 15
June 19-23 2007

Analysis of rRNA Dynamics as Inferred from Cryo-EM of Ribosomal Complexes

We are investigating various steps of mRNA-tRNA translocation during protein synthesis by cryo-EM of stalled ribosome complexes, followed by single-particle reconstruction and fitting x-ray coordinates into the resulting ~10Å density maps by real-space refinement [1]. Stalling is achieved by the addition of antibiotics or GTP nonhydrolyzable analogs. In this way, we obtain ?quasi-atomic? maps, which are plausible interpretations of the complex in the observed conformation. Comparison of such maps provides insights into the dynamics of the ribosome, especially of the ribosomal RNA.

One of the dynamical steps that appears to be required for translocation is the ratchet motion, a rotation of the small subunit relative to the large subunit [2]. It is accompanied by internal changes of both subunits in conformation. Interestingly, during protein translation, several other subprocesses unrelated to translocation appear to make use of the same intersubunit motion, namely initiation (binding of IF2 [3]), termination (binding of RF3 [4,5]), and recycling (binding of RRF [6]). A plausible hypothesis which has gained strength recently (e.g., [7]) is that the very architecture of the ribosome lends itself to the existence of two ?macro-states?, and that only a small activation barrier has to be overcome to go from one to the other.

The questions raised by these findings are (i) what is the specific purpose of the ratchet motion in each of the subprocesses, and (ii) are the conformations in the macro-states triggered by different factor binding identical, or do they differ in a way related to the functional requirements in the specific subprocess? We investigated these questions by comparing the 3D ribosomal structures (i.e., the quasi-atomic maps) directly and by making use of a newly developed RNA secondary structure mapping tool, coloRNA [8].

References and Footnotes
  1. Gao, H., Sengupta, J., Valle, M., Korostelev, A., Eswar, N., Stagg, S.M., Van Roey, P., Agrawal, R.K., Harvey, S.C., Sali, A., Chapman, M.S., and Frank, J. Cell 113, 789-801 (2003).
  2. Frank,J., and Agrawal, R.K. Nature 406, 318-322 (2000).
  3. Allen, G.S., Zavialov, A., Gursky, R., Ehrenberg, M., and Frank. J. Cell 121, 703-712 (2005).
  4. Klaholz, B.P., Myasnikov, A.G., and van Heel, M. Nature 427, 862-865 (2004).
  5. Gao, H., Zhou, Z., Rawat, U., Huang, C., Bouakaz, L., Wang, C., Cheng, Z., Liu,Y., Zavialov, A., Gursky, R., Sanyal, S., Ehrenberg, M., Frank, J., and Song, H. Cell, in press. (2007).
  6. Gao, N., Zavialov, A., Li,W., Sengupta, J., Valle, M., Gursky, R., Ehrenberg, M., and Frank, J. Mol. Cell 18, 663-674 (2005).
  7. Tama, F., Valle, M., Frank, J., and Brooks, C.L., III. Proc. Natl. Acad. Sci. USA 100, 9319-9323 (2003).
  8. LeBarron, J., Mitra, K., and Frank, J. J. Struct. Biol. 157, 262-270 (2007).

Joachim Frank1, 2
Jamie LeBarron3, and
Haixiao Gao3

Wadsworth Center,
Empire State Plaza,
Albany, New York 12201-0509

Dept of Biomedical Sci.2,
SUNY Albany,
Wadsworth Center

Wadsworth Center3

phone: 518 474-7002
fax: 518 486-2191
Email: joachim@wadsworth.org