Purification and properties of nucleoside triphosphate-adenosine monophosphate transphosphorylase from beef heart mitochondria

Mitochondrial GTP‐AMP Phosphotransferase.

“…Since UTP gives only slight activity and CTP and ATP do not serve as phosphate donors in our enzymatic preparation, it appears likely that at least one other enzyme, capable of catalyzing the interaction with nucleotide phosphates, was present in the preparation reported by Albrecht. The Michaelis constants reported in Table 3 agree within about 4-fold of values reported by Albrecht [5] with the exception of K, for GDP which is 30-fold higher. Part of these differences may be due to dependence on coupled enzymes in assays of the earlier work.…”

“…GTP-AMP phosphotransferase from beef heart mitochondria was reported by Albrecht [5] as having a molecular weight of 52000, optimum activity at pH 8.5, specific requirement for divalent cations and specific for AMP as phosphate acceptor and unspecific for the phosphate donor since GTP, ITP and to a lesser extent UTP, CTP and ATP could fulfill this role.…”

Mitochondrial GTP‐AMP Phosphotransferase

“…On the basis of 32P-incorporation studies with rat liver mitochondria, Tokumitsu and Ui [I91 suggested that the enzyme is not exposed to the outside of the inner membrane. Methods of purification resorting to sonication after removal of the outer membrane by washing mitochondria with dilute phosphate buffer [5] or by freezing and thawing as in the present purification procedure indicate the enzyme is in the matrix or bound to the inside of the inner mitochondrial membrane. Since methods of extracting GTP-AMP phosphotransferase are relatively mild, the enzyme if membrane-bound is not tightly bound to the inner membrane.…”

“…The molecular weight was found to be 26000 and the isoelectric point to be 9.8. Amino acid analysis showed 21 aspartic acid or asparagine, 19 threonine, 12 serine, 26 glutamic acid or glutamine, 15 proline, 16 glycine, 14 alanine, 15 valine, 4 methionine, 12 isoleucine, 28 leucine, 7 tyrosine, 7 phenylalanine, 5 histidine, 14 lysine, 16 arginine, 2 tryptophan, no -SSbonds or free -SH. Guanosine(5')pentaphospho(5')adenosine is a very strong inhibitor similar to adenosine(5')pentaphospho(5')adenosine as an inhibitor of cytosolic adenylate kinase.…”

“…In studies of the reaction sequence of substrate level phosphorylation in rat liver mitochondria, Heldt and Schwalbach [4] concluded that a GTP-AMP phosphotransferase is involved. More recently Albrecht [5] purified the (1) NTP-AMP transphosphorylase 41-fold from beef heart mitochondria and reported properties of the enzyme. After Khoo and Russell [6] had shown that the two isozymes of adenylate kinase in human tissue designated AKI and AK;?…”