Release factors 2 from Escherichia coli and Thermus thermophilus: structural, spectroscopic and microcalorimetric studies.
Prokaryotic class I release factors (RFs) respond to mRNA stop codons and terminate protein synthesis. They interact with the ribosomal decoding site and the peptidyl-transferase centre bridging these 75 A distant ribosomal centres. For this an elongated RF conformation, with partially unfolded core domains II.III.IV is required, which contrasts the known compact RF crystal structures. The crystal structure of Thermus thermophilus RF2 was determined and compared with solution structure of T. thermophilus and Escherichia coli RF2 by microcalorimetry, circular dichroism spectroscopy and small angle X-ray scattering. The structure of T. thermophilus RF2 in solution at 20 degrees C is predominantly compact like the crystal structure. Thermodynamic analysis point to an initial melting of domain I, which is independent from the melting of the core. The core domains II.III.IV melt cooperatively at the respective physiological temperatures for T. thermophilus and E. coli. Thermodynamic analyses and the X-ray scattering results for T. thermophilus RF2 in solution suggest that the compact conformation of RF2 resembles a physiological state in absence of the ribosome.
Zoldák
Gabriel
G
Department of Biochemistry, P. J. Safárik University Kosice, Slovakia.
Redecke
Lars
L
Svergun
Dmitri I
DI
Konarev
Peter V
PV
Voertler
C Stefan
CS
Dobbek
Holger
H
Sedlák
Erik
E
Sprinzl
Mathias
M
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry* / Q000302:isolation & purification; D002152:Calorimetry, Differential Scanning; D002942:Circular Dichroism; D003001:Cloning, Molecular; D018360:Crystallography, X-Ray; D029968:Escherichia coli Proteins / Q000737:chemistry; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010454:Peptide Termination Factors / Q000737:chemistry* / Q000302:isolation & purification; D011489:Protein Denaturation; D053838:Scattering, Small Angle; D016963:Thermus thermophilus; D014961:X-Ray Diffraction
D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D010454:Peptide Termination Factors; C489006:prfB protein, E coli