Isolation, characterization, and in vitro expression of a cDNA that encodes the kidney isoenzyme of the mitochondrial glutaminase.

A cDNA that encodes the kidney isoenzyme of the mitochondrial glutaminase (pGA) was generated by recombination of two cDNAs that were isolated from a random-primed rat brain lambda gt11 library. pGA encodes 674 amino acids which includes an N-terminal sequence of 16 residues that should form an amphipathic helix, typical of a mitochondrial targeting sequence. Residues 73-90 correspond to the N-terminal sequence of the more abundant 65-kDa glutaminase peptide. In vitro transcription and translation of pGA yields a 72-kDa peptide that is immunoprecipitated with glutaminase-specific antibodies. Incubation of the glutaminase precursor with isolated mitochondria yields the 68- and 65-kDa peptides that are characteristic of the mature glutaminase. Thus, the two mature glutaminase peptides are synthesized from a single precursor. The complete 3' nontranslated region of the GA mRNA was characterized by sequencing a GA cDNA (pGA12) that was isolated from an oligo(dT)-primed rat kidney lambda gt10 library. This segment contains numerous AU-rich regions, four potential stem-loop structures, and a 48 base pair repeat of CA dinucleotides. Such domains may contribute to the increased stability of the GA mRNA that occurs in response to metabolic acidosis.