ABSTRACT
Insulin-degrading neutral proteinase with molecular weight of 70 kDa was partly purified from the rat liver and erythrocyte plasma membranes. Incubation of membranes with [gamma-32P]ATP resulted in the enzyme phosphorylation. Intensity of this process greatly increased in the presence of insulin (100 microU/ml), and correlated with the elevation of the insulin-degrading activity in proteinase. Ca2+, Mn2+, dithiothreitol, cysteine were shown to have a stimulatory effect on insulin degradation; p-chloromercuribenzoate significantly repressed this process. Phenylmethylsulphonyl fluoride and soybean trypsin inhibitor did not affect the activity of the proteinase. It was concluded that the investigated enzyme was a calpain and may participate in the mechanism of insulin action.
Subject(s)
Endopeptidases/metabolism , Erythrocyte Membrane/enzymology , Insulin/metabolism , Liver/enzymology , Animals , Calcium/pharmacology , Cell Membrane/enzymology , Chromatography, Liquid , Cysteine/pharmacology , Dithiothreitol/pharmacology , Endopeptidases/isolation & purification , Hydrogen-Ion Concentration , Liver/cytology , Manganese/pharmacology , Proteins/analysis , RatsABSTRACT
It is established that insulin enhances the ability of the loach liver plasma membranes to phosphorylate lactate dehydrogenase. In the case of insulin-treated plasma membranes the amount of incorporated 32P is more than 4 times higher than that of the basal level. It is concluded that insulin-stimulated plasma membrane-dependent phosphorylation of the enzyme is one of the possible molecular mechanisms of hormone action on intracellular metabolism.
Subject(s)
Insulin/pharmacology , L-Lactate Dehydrogenase/metabolism , Liver/metabolism , Animals , Cell Membrane/drug effects , Cell Membrane/metabolism , Cypriniformes , In Vitro Techniques , Liver/cytology , Liver/drug effects , PhosphorylationABSTRACT
The data concerning proteolytic enzymes in human and animal malignant tumours are reviewed. The activity of these proteinases, its changes in the course of the disease, intracellular localization and secretion into the extracellular space and blood are discussed. The role of proteolytic enzymes including the role of plasminogen activator in tumour progress and during the metastasis development is considered.
Subject(s)
Neoplasms, Experimental/enzymology , Neoplasms/enzymology , Peptide Hydrolases/metabolism , Animals , Cattle , Cell Membrane/enzymology , Cell Transformation, Neoplastic/metabolism , Enzyme Activation , Humans , Mice , Neoplasm Metastasis , Neoplasm Proteins/metabolism , Neoplasms/immunology , Neoplasms, Experimental/immunology , Peptide Hydrolases/immunology , Plasminogen Activators/physiology , RatsABSTRACT
The action and some properties of cathepsin D, partly purified from unfertilized loach eggs, embryos and skeletal muscles were determined. The enzyme from embryo cells displays the activity maximum at pH 3.0 and pH 4.8 while enzyme from skeletal muscles--only at pH 3.0. Cathepsin D purified from all three sources splits actively hemoglobin, albumin, alpha-glycerophosphate dehydrogenase, pyruvate kinase and practically does not influence casein, hexokinase, glucose-6-phosphate dehydrogenase. The enzyme is comparatively thermolabile and its activity decreases in the presence of thiol compounds. The main part of cathepsin D in skeletal muscle cells and in embryo cells is precipitated after differential centrifugation of homogenates (25000 g; 60 min).
Subject(s)
Cathepsin D/analysis , Embryo, Nonmammalian/enzymology , Muscles/enzymology , Ovum/enzymology , Acid Phosphatase/analysis , Acid Phosphatase/isolation & purification , Animals , Cathepsin D/isolation & purification , Female , Fishes , Substrate SpecificityABSTRACT
Loach eggs are stated to possess some quantities of insulin or insulin-like substances with a positive reaction to antiinsulin antiserum. The level of these components is five times as high 30 minutes after fertilization, and does not change for the next 20 hours of the embryo development. No 17-oxycorticosteroids were detected in loach eggs and embryos during first four hours of the development; they appeared only at the blastula stage and at the state of gastrula their level increases considerably. It is supposed that insulin or insulin-like substances participate in metabolism regulation and cell differentiation of fish embryos during early stages of embryogenesis.
Subject(s)
17-Hydroxycorticosteroids/analysis , Embryo, Nonmammalian/analysis , Fishes/embryology , Insulin/analysis , Ovum/analysis , Animals , RadioimmunoassayABSTRACT
The content of glycogen and glucose, as well as aldolase, phosphofructokinase, phosphoglucomutase, glucose-6-phosphatase and fructose-1,6-diphosphatase activities in liver tissue and the same activities in skeletal muscle of sheep were determined under the influence of prolonged addition of carboxyline separately and in combination with methionine, diammonium phosphate and potassium iodine to their diet. It is established that under the influence of carboxyline the glycogen content as well as aldolase and fructose-1,6-diphosphatase activities rise significantly in the liver of the tested animals. In the skeletal muscle only aldolase activity increases.
Subject(s)
Bicarbonates/pharmacology , Carbohydrate Metabolism , Liver/metabolism , Metals/pharmacology , Muscles/metabolism , Quaternary Ammonium Compounds/pharmacology , Sodium Bicarbonate , Animals , Drug Combinations/pharmacology , Fructose-Bisphosphatase/metabolism , Fructose-Bisphosphate Aldolase/metabolism , Glucose/metabolism , Glucose-6-Phosphatase/metabolism , Glycogen/metabolism , Liver/drug effects , Liver Glycogen/metabolism , Muscles/drug effects , Phosphofructokinase-1/metabolism , Phosphoglucomutase/metabolism , SheepABSTRACT
The activities of hexokinase, glucokinase, phosphofructokinase, glucose-6-phosphate dehydrogenase, glucose-6-phosphatase, and fructose-1,6-diphosphatase were determined in loach embryos developed in solutions of insulin, hydrocortisone, estrone and thyroxin at different stages of embryogenesis. Glucokinase and fructose-1,6-diphosphatase activties are shown not to change markedly under the influence of the above-mentioned hormones. During some periods of early development the hexokinase activity is inhibited by insulin, estrone and thyroxin. The glucose-6-phosphate dehydrogenase activity is suppressed by each of the used hormones at all the stages of early embryogenesis while the glocose-6-phosphatase activity decreased only under the influence of insulin at the cleavage, blastula and gastrula stages. Insulin increased the activity of phosphofructokinase at the cleavage, blastula and early gastrula stages and hydrocortisone, estrone and thyroxine during certain periods of these stages. From middle gastrula two last hormones decreased the phosphofructokinase activity in the loach embryos.