Transgenic livers expressing mitochondrial and cytosolic CK: mitochondrial CK modulates free ADP levels

Abstract

Askenasy, Nadir, and Alan P. Koretsky. Transgenic livers expressing mitochondrial and cytosolic CK: mitochondrial CK modulates free ADP levels. Am J Physiol Cell Physiol 282: C338-C346, 2002. First published October 3, 2001 10.1152/ajpcell.00404.2001.-The function of creatine kinase (CK) and its effect on phosphorus metabolites was studied in livers of transgenic mice expressing human ubiquitous mitochondrial CK (CK-Mit) and rat brain CK (CK-B) isoenzymes and their combination.31 P NMR spectroscopy and saturation transfer were recorded in livers of anesthetized mice to measure high-energy phosphates and hepatic CK activity. CK reaction velocity was related to total enzyme activity irrespective of the isoenzyme expressed, and it increased with increasing concentrations of creatine (Cr). The fluxes mediated by both isoenzymes in both directions (phosphocreatine or ATP synthesis) were equal. Over a 20-fold increase in CK-Mit activity (28-560 mol ⅐ g wet wt Ϫ1 ⅐ min Ϫ1 ), the fraction of phosphorylated Cr increased 1.6-fold. Hepatic free ADP concentrations calculated by assuming equilibrium of the CK-catalyzed reaction in vivo decreased from 84 Ϯ 9 to 38 Ϯ 4 nmol/g wet wt. Calculated free ADP levels in mice expressing high levels of CK-B (920-1,635 mol ⅐ g wet wt Ϫ1 ⅐ min Ϫ1 ) were 52 Ϯ 6 nmol/g wet wt. Mice expressing both isoenzymes had calculated free ADP levels of 36 Ϯ 4 nmol/g wet wt. These findings indicate that CK-Mit catalyzes its reaction equally well in both directions and can lower hepatic apparent free ADP concentrations. mitochondrial creatine kinase; cytosolic creatine kinase; phosphorus-31 nuclear magnetic resonance spectroscopy; magnetization transfer; liver adenosine 5Ј-diphosphate concentration

31P Saturation Transfer Spectroscopy Predicts Differential Intracellular Macromolecular Association of ATP and ADP in Skeletal Muscle

“…2 for the mutant and WT mice indicate that they are proportional to CK activity, reflecting its concentration. A linear relation between k CK,for and CK activity (V max ) has been noticed previously with different expression levels of the M-CK isoform (35). It is inherent in the present approach that saturation of the spin population of bound ADP is not possible.…”

Studying enzymes by in vivo 13C magnetic resonance spectroscopy

“…The ability of noninvasively extracting information from specific enzymes using in vivo MRS is highly significant and has generated a great deal of enthusiasm [14–21]. In particular, creatine kinase-catalyzed magnetization transfer effect has been demonstrated to be a useful magnetic resonance reporter of gene expression [22]. Obviously, it would be highly desirable if more enzymes were accessible to in vivo MRS-based magnetization transfer spectroscopy methods.…”

Impaired Intracellular Energetic Communication in Muscles from Creatine Kinase and Adenylate Kinase (M-CK/AK1) Double Knock-out Mice

“…Skeletal muscle also contains an additional mitochondrial CK isoform (ScCKmit), which amounts to 1-10% of the total CK activity depending on the type of muscle fiber (26,27). This CK member associates and functionally interacts with the adenine nucleotide translocator and voltage-dependent anion channel in the mitochondrial inner and outer membrane (28 -30), providing an efficient ATP export and metabolic signal reception pathway (31).…”