Several recent studies have demonstrated the presence of creatine kinase and of phosphorylcreatine in a variety of cells besides striated muscle and brain cells. The total creatine kinase and phosphagen levels in these cells encompass a wide range of values. The available data are collected in this article to demonstrate that the variation of the enzyme and phosphagen concentrations is not random but that the two are interrelated. With both the major isoenzymes of creatine kinase, namely the muscle type and the brain type, the basal levels of phosphorylcreatine follow closely the cellular creatine kinase levels. A hypothesis is presented in which the enzyme itself is the major determinant of phosphorylcreatine content by virtue of its ability to act as an intracellular binding protein for creatine derived from extracellular fluid, and also for cellular ADP. The proposed mechanism further predicts that in cells that contain high levels of actin and thus sequester the cytoplasmic free ADP (e.g. most muscle cells), a high level of creatine kinase can effectively regulate the myokinase reaction by its ability to bind ADP. The net effect of such regulation is to conserve the adenine nucleotide pools in the cell. The evolutionary advantage of these two regulatory functions of creatine kinase in terms of energy conservation is discussed.
In the Mexican axolotl Ambystoma mexicanum recessive mutant gene c, by way of abnormal inductive processes from surrounding tissues, results in an absence of embryonic heart function. The lack of contractions in mutant heart cells apparently results from their inability to form normally organized myofibrils, even though a few actin-like (60-A) and myosin-like (150-A) filaments are present. Amorphous "proteinaceous" collections are often visible. In the present study, heavy meremyosin (HMM) treatment of mutant heart tissue greatly increases the number of thin filaments and decorates them in the usual fashion, confirming that they are actin. The amorphous collections disappear with the addition of HMM. In addition, an analysis of the constituent proteins of normal and mutant embryonic hearts and other tissues is made by sodium dodecyl sulfate (SDS) gel electrophoresis. These experiments are in full agreement with the morphological and HMM binding studies. The gels show distinct 42,000-dalton bands for both normal and mutant hearts, supporting the nresence of normal actin. During early developmental stages (Harrison's stage 34) the cardiac tissues in normal and mutant siblings have indistinguishable banding patterns, but with increasing development several differences appear. Myosin heavy chain (200,000 daltons) increases substantially in normal hearts during development but very little in mutants. Even so, the quantity of 200,000-dalton protein in mutant hearts is significantly more than in any of the nonmuscle tissues studied (i.e. gut, liver, brain). Unlike normal hearts, the mutant hearts lack a prominent 34,000-dalton band, indicating that if mutants
High levels of phosphocreatine, a compound known to serve as an intracellular energy reserve, were found in the fluid contained in seminal vesicle glands. The concentrations of phosphocreatine in the extracellular fluid in the mouse and rat were found to be 5.6 ± 1.6 and 2.2 ± 0.8 ,umol/g, respectively, which are higher than the intracellular levels reported for smooth muscles. The creatine concentrations in the seminal vesicular fluid from these two species were 22.8 ± 3.1 and 13.0 ± 5.3 ,umol/g, respectively. These creatine levels are approximately 100 and 65 times higher than the creatine levels in mammalian blood. Smaller amounts of ATP (phosphocreatine/ATP ratio of 20-40) and traces of ADP were also found. Comparison of the pattern of distribution of macromolecules (proteins and DNA) with the distribution of phosphocreatine between the cells and the fluid of the seminal vesicle indicates that cell lysis did not account for the phosphocreatine in the seminal vesicle fluid. Rather, the available evidence strongly suggests that this high-energy compound is actively secreted. We found that in the testes, the sperm are exposed to the highest known creatine concentration in any mammalian tissue studied. Based on these results and other recent reports, we propose that the extracellular phosphocreatine, ATP, and creatine are involved in sperm metabolism.Phosphocreatine (PCr), a guanidinophosphate, was first discovered in skeletal muscle (1,2). It is believed to serve as an energy reserve by virtue of its ability to phosphorylate ADP, leading to the production of ATP and creatine (Cr). This reversible phosphoryl transfer, mediated by creatine kinase (CK; ATP:creatine N-phosphotransferase, EC 2.7.3.2), can be represented as follows. CK/PCr-mediated energy modulator systems have since been demonstrated in several other cell types, including brain (3, 4), smooth muscle (5-7), mammalian preimplantation embryo (8), and spermatozoa (9, 10), and in the mitotic spindle of proliferating animal cells (11). In addition to an ATP-buffering role, reaction 1 has been suggested to act as an intracellular energy-transport system (12). Strong experimental evidence for such an energy-channeling role for PCr in sea urchin sperm has been produced (10). PCr and Cr in higher organisms are considered to be dead-end metabolites in that the CK-mediated reversible transfer of phosphate groups is the only known enzyme reaction in which PCr and Cr participate. However, a small portion of the total body creatine (PCr + Cr) is converted by a nonenzymatic reaction to the anhydride, creatinine, which can readily cross cell membranes and is the excretory product of PCr and Cr (13).It has been shown (14) MATERIALS AND METHODS Seminal Vesicles and Vesicular Fluid. Seminal vesicles were obtained from Swiss Webster mice (8 weeks old) or from Sprague-Dawley rats (10-12 weeks old). The animals were acclimated to a standard laboratory chow and to a 12-hr light/dark cycle for 5-7 days. They were killed by inhalation of CO2 in a large desiccator. T...
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