Possible participation of calmodulin in stimulation of leucine transport by concanavalin A in human lymphocytes

Stimulation of leucine uptake by addition of concanavalin A, mediated by increase of intracellular free Ca2+ concentration [( Ca2+]), in lymphocytes (Mitsumoto, Y., Sato, K. and Mohri, T. (1988) Biochim. Biophys. Acta 968, 353-358) was abolished by N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide (W-7) and chlorpromazine, which inhibited membrane hyperpolarization induced by the mitogen. Quinine (0.5-1 mM) completely inhibited the concanavalin A-induced hyperpolarization and extensively inhibited the induced stimulation of leucine uptake. Based on these results, we suggest that the stimulation of leucine uptake by concanavalin A is largely due to activation of the Ca2+-dependent K+ channel which reinforces negative potential of the plasma membrane and is regulated by calmodulin.     

The effects of insulin on choline kinase activity in cultured rat liver cells

The effect of insulin on phosphatidylcholine biosynthesis in cultured rat liver cells was assessed by measuring changes in the activity of the first enzyme in the choline pathway of phosphatidylcholine biosynthesis, choline kinase (ATP: cholinephosphortransferase, EC 2.7.1.32), in the presence or absence of the hormone. Choline kinase specific activity in liver cells incubated for 18 hours in the presence of 10^?7M insulin increased two-fold from 3.4 ± 0.3 nmoles phosphorylcholine formed/min/mg protein to 7.5 ± 0.6 nmoles/min/mg protein. This effect was dose dependent and reversed by the addition of actinomycin D and cycloheximide. It is concluded that the increase in specific activity is due to synthesis of new enzyme rather than activation of existing enzyme.     

Reasons for disorders of fatty acid oxidation in isolated heart mitochondria during ischemia

The influence of malate and cytochrome c on fatty acid oxidation under control and ischemic conditions was investigated. In the medium without malate, cytochrome did not make fatty acid oxidation decreased during ischemia return to normal. Oxidation in the media containing malate and cytochrome did not differ from control only when it was measured after preliminary oxidation of endogenous substrates. The ratio of palmitoyl-CoA and palmitoyl carnitine to the respiration rates at state 3 was unchanged at 60 min ischemia. Apparently, no changes in carnitine acyltransferase playing a role in oxidation of palmitoyl-CoA took place. Thus, the decrease of fatty acid oxidation at early periods of ischemia is largely caused by a reduction in the content of cytochrome c and intermediates of Krebs cycle in the mitochondria.     

Mechanistic studies on carboxypeptidase a from goat pancreas — part II: Evidence for carboxyl group

Studies with substrate analogues and the pH optimum indicated the involvement of carboxyl group in the active site of goat carboxypeptidase A. Chemical modification of the enzyme with 1-cyclohexyl-3-(2-morpholinoethyl) carbodiimide methoI -p-toluene sulphonate, a carboxyl specific reagent, led to loss of both esterase and peptidase activities. Protection studies showed that this carboxyl group was in the active site and was protected by Βp-phenylpropionic acid and glycyl-L-tyrosine. Kinetic studies also confirmed the involvement of carboxylic group because the enzyme modification with water soluble carbodiimide was a two step reaction which excluded the possibility of tyrosine or lysine which are known to give a one step reaction with this reagent