Interactions between calmodulin and immobilized phenothiazines.

The ability of a number of antipsychotic drugs such as phenothiazines to bind to calmodulin with high affinity in a calcium-dependent manner was applied to the study of the nature of their interactions with calmodulin. Thus, a series of phenothiazine derivatives and analogues were immobilized on agarose and examined for their binding characteristics to calmodulin. The binding of calmodulin to fluphenazine, perphenazine and 7-aminotriflupromazine involved on the one hand non-specific electrostatic interactions which are abolished by increasing the eluent salt concentration, and on the other hand, Ca2+-dependent interactions which are reversed by EGTA addition.

Protein kinases and their protein substrates in free messenger ribonucleoprotein particles and polysomes from mouse plasmacytoma cells.

In free messenger ribonucleoprotein particles (mRNP) and polysomes from plasmacytoma cells, a phosphorylated protein/protein kinase system has been characterized by a combination of oligo(dT)-cellulose chromatography and CsCl isopycnic gradient centrifugation. The presence phosphorylated in vivo has been detected in both types of particles. Endogenous protein phosphorylation occurs in vitro by particle-associated cAMP-independent protein kinase(s) using [gamma-32P]ATP and [gamma-32P]GTP.

In vitro translation studies of the cytoplasmic nonpolysomal particles containing messenger RNA.

We analyzed the translational capacity of different kinds of free cytoplasmic messenger ribonucleoprotein complexes (free mRNP) in a Hela cell cell free system. Native free mRNP are not translated although free mRNP washed with 0.5 M KC1 can direct polypeptide synthesis.

A rapid direct assay of 3'-5' cyclic nucleotide phosphodiesterase activity using chromatography on immobilized acriflavin.

A chromatographic method using immobilized acriflavin has been developed for the separation of unreacted cyclic nucleotides from their corresponding 5'-nucleotides, in a direct assay of 3'-5' cyclic nucleotide phosphodiesterase using [3H]-cyclic nucleotides as substrate. The method based on the so-called charge-transfer overlap recognition between flavin and indol rings, provides a rapid (15-20 min) and sensitive elution of [3H]-5'nucleotides with high recovery (up to 98%) and low blanks, while [3H]-cyclic nucleotides are retarded on the column. By this method, the formation of some secondary products by purine metabolizing enzymes such as 5'-nucleotidases, nucleosidases and/or deaminases is taken into account, using [14C]-5'-AMP thus allowing an accurate determination of phosphodiesterase activity in any preparations.

A new chromatographic method using immobilized acriflavin for measuring cyclic AMP in cells prelabeled with radioactive adenine.

A method using the principle of charge-transfer chromatography has been developed for the determination of cyclic AMP levels in intact prelabeled cells. The ATP pool was prelabeled by incubating the cells in the presence of radioactive adenine. The cyclic AMP formed from ATP was extracted with HC10(4) and separated from adenine and other adenosine-related nucleotides by chromatography on acriflavin-Sephadex G-25.

mRNP proteins, initiation factors and phosphorylation.

Information has been collected to stimulate interest regarding the nature and the possible role of mRNP protein phosphorylation in a cytoplasmic control mechanism for protein synthesis. It does not imply a direct relationship between mRNP protein and initiation factors. These proteins have some properties in common (e.

Purification of a soluble ATPase from rat liver mitochondria by AMP-Sepharose affinity chromatography.

ATPase (ATP phosphohydrolase, EC 3.6.1.

[Ribonucleoproteins containing heterogeneous nuclear and messenger cytoplasmic RNA. Characteristics, structure and relations (author's transl)].

Following the study of Spirin, many authors have shown that cytoplasmic messenger RNA and heterogeneous nuclear RNA are complexed with specific proteins to form ribonucleoprotein particles (RNP). These RNP are heterogeneous in size and present a high protein to RNA ratio. Different observations suggest a polymeric structure for nuclear ribonucleoproteins but their protein composition is more clearly complex than that of cytoplasmic ribonucleoproteins.

Characterization of a protein kinase-phosphoprotein system in free cytoplasmic ribonucleoprotein particles of plasma cell tumours.

A protein kinase, associated with free cytoplasmic ribonucleoprotein particles (free dRNP) has been purified from mouse plasma cell tumours. This protein kinase is able to phosphorylate in vitro endogenous protein from free dRNP. Some characteristics of this protein kinase have been studied.

A plasmocytoma ribosome-associated protein kinase which phosphorylates a specific protein of the ribosomal KCl wash.

One ribosomal protein kinase activity and 3 soluble protein kinase activities have been identified in plasma cell tumors by DEAE-cellulose chromatography. We have shown phosphorylation in vivo and in vitro of a protein fraction from the ribosomal KCl wash which we have termed 'PPx fraction'. Phosphorylation of this protein fraction has been obtained in vitro with the ribosome-associated protein kinase.

[Demonstration of isoenzymes of human serum N-acetyl-beta-D-hexosaminidases, during pregnancy].

The N-acetyl-beta-D-hexosaminidase activity (EC 3.2.1.