In vitro mutagenesis of the mitochondrial leucyl tRNA synthetase of Saccharomyces cerevisiae shows that the suppressor activity of the mutant proteins is related to the splicing function of the wild-type protein.
The NAM2 gene of Saccharomyces cerevisiae encodes the mitochondrial leucyl tRNA synthetase (mLRS), which is necessary for the excision of the fourth intron of the mitochondrial cytb gene (bI4) and the fourth intron of the mitochondrial coxI gene (aI4), as well as for mitochondrial protein synthesis. Some dominant mutant alleles of the gene are able to suppress mutations that inactivate the bI4 maturase, which is essential for the excision of the introns aI4 and bI4. Here we report mutagenesis studies which focus on the splicing and suppressor functions of the protein. Small deletions in the C-terminal region of the protein preferentially reduce the splicing, but not the synthetase activity; and all the C-terminal deletions tested abolish the suppressor activity. Mutations which increase the volume of the residue at position 240 in the wild-type mLRS without introducing a charge, lead to a suppressor activity. The mutant 238C, which is located in the suppressor region, has a reduced synthetase activity and no detectable splicing activity. These data show that the splicing and suppressor functions are linked and that the suppressor activity of the mutant alleles results from a modification of the wild-type splicing activity.
Li
G Y
GY
Centre de Génétique Moléculaire, Laboratoire propre du CNRS associé á I'Université Pierre et Marie Curie, Gif-sur-Yvette, France.
Bécam
A M
AM
Slonimski
P P
PP
Herbert
C J
CJ
D004722:Endoribonucleases / Q000235:genetics; D005800:Genes, Fungal; D007935:Leucine-tRNA Ligase / Q000235:genetics*; D008928:Mitochondria / Q000201:enzymology; D009713:Nucleotidyltransferases / Q000235:genetics; D012326:RNA Splicing; D012331:RNA, Fungal / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics*; D013489:Suppression, Genetic
D012331:RNA, Fungal; D009713:Nucleotidyltransferases; C034038:mRNA maturase; D004722:Endoribonucleases; D007935:Leucine-tRNA Ligase