MITOCHONDRIAL ASPARAGINYL-TRANSFER-RNA SYNTHETASE IS ENCODED BY THE YEAST NUCLEAR GENE YCR24C

One of the open reading frames located on yeast Saccharomyces cerevisi ae chromosome III, YCR24c, appeared to code for a protein of unknown f unction, but the predicted sequence showed similarity with asparaginyl -tRNA synthetase from Escherichia coli, with 38% amino acid identity. There is a putative mitochondrial targeting signal at the N-terminus o f the YCR24c product. Northern blot analysis of total RNA from a wild- type strain Sigma 1278b confirmed that YCR24c was transcribed. Disrupt ion of the chromosomal copy of YCR24c in a respiratory-competent haplo id cell induced a petite phenotype, but did not affect cell viability. This respiratory-defective phenotype is typical for a mutation in a n uclear gene that induces a non-functional mitochondrial protein synthe sis system. The protein encoded by YCR24c was expressed in Escherichia coli in a histidine tagged form and tid isolated. The enzyme aminoacy lated unfractionated Escherichia coli tRNA with asparagine. These resu lts identified YCR24c as the structural gene for yeast mitochondrial a sparaginyl-tRNA synthetase.