In vivo and in vitro analysis of structure-function relationships in ribosomal protein L25 from Saccharomyces cerevisiae.

摘要: We have developed a combination of in vivo and vitro methods which allows us to determine the effect practically every structural change, deletions as well point mutations, on various biological functions ribosomal protein (r-protein). used this approach delineate functional domains r-protein L25 from Saccharomyces cerevisiae. By analysis intracellular distribution fusion proteins carrying portions linked Escherichia coli β-galactosidase we traced nuclear localization signal(s) region encompassing N-terminal 61 amino acids protein. On other hand, using prepared fragments located domain responsible for its specific binding 26S rRNA between 135. In order be able analyze mutations ribosome biogenesis function constructed mutant yeast strain chromosomal gene is placed under control inducible GAL promoter. Since unable grow glucose carbon source must essential cell viability. Growth can restored by introduction wild-type plasmid, demonstrating that these conditions cells are dependent upon an extrachromosomally supplied copy gene.