Involvement of the 3' side of the anticodon loop of yeast tRNATyr in messenger-free binding to ribosomes. An electron-spin resonance study.

Abstract

Electron-spin resonance (ESR) spectra of a nitroxide spin-label attached to residue i6A-37 of yeast tRNATyr were measured in complexes of deacylated tRNATyr with Escherichia coli ribosomes. A Scatchard plot, obtained in the absence of mRNA, indicated strong binding with an association constant of 1 X 10(7) l X mol-1, suggesting the P-site binding. The ESR spectrum of free tRNATyr, characteristic for a rapidly tumbling nitroxide, changes to a spectrum with extensively broadened lines in the ribosome-tRNA complex. The original spectrum can be restored upon long incubations of the complex with an excess of extraneous tRNA. ESR spectra suggest that the spin-label motion is drastically perturbed though not completely blocked in the ribosome-tRNATyr complex. Since ESR spectra of a spin-label attached to the opposite, i.e. 5', side of the anticodon loop are only slightly perturbed by the messenger-free binding to ribosomes [Rodriguez et al. (1980) J. Biol. Chem. 255, 8116-8120], it is concluded that the two sides of the anticodon loop face entirely different environments when bound to the P site, the 3' side being oriented towards the surface of the ribosome, and the other side towards its environment or a large cavity.