Phospholipase of rat intestine: mode of action]

Phospholipase activities of rat intestinal mucosa homogenate have been determined from lysophosphatidylcholines [14C] and phosphatidylcholines [-3H-14C]. In the presence of phosphatidylcholines, at pH 6.5, the homogenate has a phospholipase B activity. At pH 8.5, a phospholipase A2 activity was shown. In the presence of lysophospatidylcholines, at pH 6.5, we notice a lysophospholipase A1 activity. A kinetic study of the reactions allows us to separate the activity B into a phospholipase A2 activity and a lysophospholipase A1 activity. Thus, it appears that the total phospholipase activity of rat intestinal mucosa would results from a phospholipase A2 activity and a lysophospholipase A1 activity.     

The localization of calcium by X-ray microanalysis in myopathic muscle fibers

An electron histochemical study was undertaken to localize calcium with ammonium oxalate precipitation technique in soleus muscle of rat in normal cases and in myopathy induced experimentally by a prolonged treatment of 2,4-dichlorophenoxyacetate (2,4-D). The calcium content of precipitates was detected by energy-dispersive X-ray microanalysis. In normal cases, the electron dense precipitates containing calcium were mainly found in the vesicles of sarcoplasmic reticulum, whereas in 2,4-D induced myopathy the deposits were shifted near the Z line into the myofibrils. Calcium, because the uptake into sarcoplasmic vesicles was inhibited by 2,4-D, could attach to other binding sites, such as to the troponin-C.A long-lasting binding of calcium might lead to a prolonged activation of the actin-myosin system.     

Transient and long-term differential modulations of branched-chain α-keto acid decarboxylase activity in hypophysectomized rats

Hypophysectomy caused a marked but transient increase in branched-chain α-keto acid decarboxylase activities in rat liver mitochondria, peaking at about nine days post-surgery. The magnitude of increase is different for each of the three branched-chain α-keto acids. The activities then fall to a new steady state in three weeks with α-ketoisovalerate decarboxylase activity within the normal range, α-keto-β-methylvalerate decarboxylase activity at twice normal, and α-ketoisocaproate decarboxylase activity decreased to a level too low for accurate measurements.     

Phosphorylation of rat kidney pyruvate kinase type L by cyclic 3',5'-AMP-dependent protein kinase.

Pyruvate kinase (ATP:pyruvate 2-O-phosphotransferase, EC 2.7.1.40) type L was partly purified from rat kidney. During the last two purification steps, the incorporation of [32P]phosphate into protein on incubation with [32P]ATP and cyclic 3',5'-AMP-dependent protein kinase was found to parallel the pyruvate kinase activity. After phosphorylation of the enzyme, a major radioactive band with a molecular weight of 57 000 was found on polyacrylamide gel electrophoresis [32P]Phosphorylserine was isolated from the kidney pyruvate kinase. Immunological identity was found between the liver and kidney pyruvate kinases type L. By autoradiography of high-voltage electropherograms after partial acid hydrolysis of the phosphorylated rat liver and kidney pyruvate kinases type L, identical results were obtained. The affinity for phosphoenolpyruvate was found to be decreased by phosphorylation of the enzyme with a change in the apparent Km from 0.15 mM to 0.35 mM. After incubation of the phosphorylated kidney pyruvate kinase with phosphatase the phosphoenolpyruvate saturation curve was found to be identical to that for the unphosphorylated enzyme. Thus, the activity of the rat kidney pyruvate kinase type L is with all probability regulated by a reversible phosphorylation-dephosphorylation reaction, thereby indicating that hormonal regulation of gluconeogenesis via cyclic AMP may be of importance in the renal cortex.