Single point mutations in domain II of the yeast mitochondrial release factor mRF-1 affect ribosome binding.
We have recently described two yeast strains that are mutated in the MRF1 gene encoding the mitochondrial release factor mRF-1. Both mutants provoke gene-specific defects in mitochondrial translational termination. In the present study we report the cloning, sequencing, as well as an analysis of residual activities of both mutant mrf1 alleles. Each allele specifies a different single amino acid substitution located one amino acid apart. The amino acid changes do not affect the level or cellular localization of the mutant proteins, since equal amounts of wild type and mutant mRF-1 were detected in the mitochondrial compartment. Over-expression of the mutant alleles in wild type and mrf1 mutant yeast strains produces a phenotype consistent with a reduced affinity of the mutant release factors for the ribosome, indicating that the mutations map in a release factor domain involved in ribosome binding. We also demonstrate that nonsense suppression caused by a mutation in the mitochondrial homolog of the E. coli small ribosomal protein S4 can be reversed by a slight over-expression of the MRF1 gene.
Pel
H J
HJ
Department of Molecular Cell Biology, University of Amsterdam, The Netherlands.
Rep
M
M
Dubbink
H J
HJ
Grivell
L A
LA
D000483:Alleles; D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D017931:DNA Primers / Q000737:chemistry; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D008928:Mitochondria / Q000737:chemistry / Q000378:metabolism; D024101:Mitochondrial Proteins; D008969:Molecular Sequence Data; D010443:Peptide Chain Termination, Translational; D017354:Point Mutation; D011993:Recombinant Fusion Proteins; D012270:Ribosomes / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013329:Structure-Activity Relationship; D014157:Transcription Factors
D017931:DNA Primers; D005656:Fungal Proteins; C071720:MTF1 protein, S cerevisiae; D024101:Mitochondrial Proteins; D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors