Mitochondrial adenylate kinase from chicken liver. Purification characterization and its cell-free synthesis.
Mitochondrial adenylate kinase has been purified 5400-fold from chicken liver extract in an overall yield of 36%. The purified enzyme has a specific activity of 810 U/mg, a molecular weight of 28 000, and the following amino acid composition: 21 aspartic acid or asparagine, 14 threonine, 17 serine, 27 glutamic acid or glutamine, 16 proline, 22 glycine, 22 alanine, 15 valine, 6 methionine, 11 isoleucine, 29 leucine, 5 tyrosine, 7 phenylalanine, 16 lysine, 7 histidine, 19 arginine, 3 half-cystine, and no tryptophan, totalling 257 residues. The purified enzyme has one disulfide bond and one sulfhydryl group. The disulfide bond is related to the active conformation of the enzyme, whereas the sulfhydryl group does not contribute to the enzyme activity. The sulfhydryl group is easily oxidized in the presence of Cu2+ resulting in the formation of dimer with about one half of the specific activity of the monomer. The enzyme is similar to porcine heart mitochondrial adenylate kinase in antigenicity but different from chicken cytosolic adenylate kinase. Mitochondrial adenylate kinase was synthesized in the mRNA-dependent rabbit reticulocyte lysate system programmed with total chicken liver RNA. The mobility in sodium dodecylsulfate gel electrophoresis of the product obtained in vitro was the same as that of the purified mitochondrial adenylate kinase. This evidence indicates that the mitochondrial adenylate kinase is synthesized as a polypeptide with a molecular weight indistinguishable from that of the mature protein.
D000263:Adenylate Kinase / Q000302:isolation & purification*; D000818:Animals; D002474:Cell-Free System; D055598:Chemical Phenomena; D002621:Chemistry; D002645:Chickens; D003600:Cytosol / Q000201:enzymology; D008930:Mitochondria, Liver / Q000201:enzymology*; D008970:Molecular Weight; D010770:Phosphotransferases / Q000302:isolation & purification*
D010770:Phosphotransferases; D000263:Adenylate Kinase