Isolation and characterization of two types of cDNA for mitochondrial adenylate kinase and their expression in Escherichia coli.
Two cDNA clones for mitochondrial adenylate kinase were isolated from a cDNA library of bovine liver poly(A)+ RNA by using synthetic oligodeoxynucleotides as probes. The clone containing a 0.9-kilobase insert had the reading frame for a 241-residue protein (AK2A), while the other clone containing a 1.6-kilobase insert had the frame for a 234-residue protein (AK2B). Nucleotide sequences of these two clones were the same in the 5' portion up to the coding sequence for the 233rd residue, but different in the remaining 3' portions. The reported amino acid sequence of mitochondrial adenylate kinase from bovine heart corresponded to AK2A. Neither AK2A nor AK2B had a cleavable NH2-terminal presequence as that found in other imported mitochondrial proteins. RNA blot analysis of poly(A)+ RNAs from bovine liver and heart revealed three species of mRNA with approximate sizes of 0.9, 1.4, and 1.7 kilobases. The 1.7- and 1.4-kilobase species were specific for AK2B, whereas the 0.9-kilobase species was specific for AK2A. In the liver, the 1.7-kilobase mRNA was more abundant, whereas in the heart the 0.9-kilobase mRNA was predominant. The 1.4-kilobase mRNA was present only in the heart. The AK2A- and AK2B-coding sequences were expressed in Escherichia coli cells under the control of trc promoter. Both the products reverted the temperature-sensitive phenotype of the adenylate kinase mutant of E. coli.
D000263:Adenylate Kinase / Q000235:genetics*; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002417:Cattle; D004247:DNA / Q000302:isolation & purification*; D004926:Escherichia coli / Q000235:genetics*; D005786:Gene Expression Regulation; D007527:Isoenzymes / Q000235:genetics; D008929:Mitochondria, Heart / Q000201:enzymology; D008930:Mitochondria, Liver / Q000201:enzymology*; D010770:Phosphotransferases / Q000235:genetics*; D010957:Plasmids; D012333:RNA, Messenger / Q000032:analysis
D007527:Isoenzymes; D012333:RNA, Messenger; D004247:DNA; D010770:Phosphotransferases; D000263:Adenylate Kinase