Crystallography of halophilic ribosome: the isolation of an internal ribonucleoprotein complex.
Crystals of 50S ribosomal subunits from Haloarcula marismortui diffracting to 2.9 A resolution were grown. Because of their large unit cells and the extremely weak diffracting power, almost all X-ray crystallographic analysis of these crystals must be performed with intense synchrotron radiation. At ambient temperature, all ribosomal crystals decay upon the first instance of X-irradiation. To overcome this severe sensitivity, procedures for data collection at cryo temperature were developed. Under these conditions the crystals can be irradiated for periods sufficient for the collection of more than one data set from an individual crystal (days or weeks) with no observable damage. They also can be stored for months, to resume interrupted measurements. To assist the interpretation of the anticipated electron density map, a specific internal nucleoprotein complex of protein HmaL1 and a stretch of H23S rRNA was isolated from the halophilic ribosome. The fragments of the 23S rRNA protected by the protein from nuclease digestion were sequenced. Alignment of the sequences of some archaebacterial L1-specific RNA fragments to the corresponding parts of eubacterial and eukaryotic rDNAs, localized the sequence identities to two distinct regions. Chimeric complexes were reconstituted with the corresponding E. coli ribosomal components, indicating a rather high homology, despite the evolution distance. A feasible secondary structure of the rRNA stretch participating in this complex was found to be compatible with the one proposed for the corresponding part in the E. coli ribosomal RNA.
Max-Planck-Laboratory for Ribosomal Structure, Hamburg, Germany.
U
U
Evers
F
F
Franceschi
N
N
Böddeker
A
A
Yonath
D001483:Base Sequence; D018360:Crystallography, X-Ray; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D012261:Ribonucleoproteins / Q000737:chemistry / Q000302:isolation & purification* / Q000648:ultrastructure; D012270:Ribosomes / Q000737:chemistry / Q000648:ultrastructure*
D046911:Macromolecular Substances; D012261:Ribonucleoproteins