Cloning and sequence analysis of a cDNA encoding porcine mitochondrial aspartate aminotransferase precursor

Abstract

The primary structure of pig mitochondrial aspartate aminotransferase (mAspATase; L-aspartate:2-oxoglutarate aminotransferase, EC 2.6.1.1) precursor was deduced from its cDNA sequence. A library of cDNA clones was constructed from pig liver poly(A)+ RNA by applying the vector/primer method of Okayama and Berg [Okayama, H. & Berg, P. (1982) Mol. Cell. Biol. 2, 161-170]. The library was screened for pig mAspATase sequences by using a mixture of eight oligodeoxyribonucleotides as a probe. The sequences of the probe were deduced from the known amino acid sequence of pig mAspATase residues 196-201. Two recombinant plasmids containing inserts of about 2500 and 2600 base pairs were selected for sequence analysis. The amino acid sequence predicted from the cDNA sequence shows that the pig mAspATase precursor consists of the mature enzyme of 401 amino acid residues and an amino-terminal segment of 29 amino acid residues called the "presequence" that contains four basic amino acid residues, no acidic residues, and no hydrophobic amino acid stretch. The sequence of this 29-amino acid mAspATase precursor segment was compared with the presequences of other mitochondrial enzymes.