Processing of proenkephalin-A in bovine chromaffin cells. Identification of natural derived fragments by N-terminal sequencing and matrix-assisted laser desorption ionization-time of flight mass spectrometry.

A large variety of proenkephalin-A-derived peptides (PEAPs) are present in bovine adrenal medulla secretory granules that are cosecreted with catecholamines upon stimulation of chromaffin cells. In the present paper, after reverse phase high performance liquid chromatography of intragranular soluble material, PEAPs were immunodetected with antisera raised against specific proenkephalin-A (PEA) sequences (PEA63-70 and PEA224-237) and analyzed by matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry. Thirty PEAPs were characterized in addition to enkephalins and whole PEA, indicating that preferential proteolytic attacks occurred at both N- and C-terminal regions. A similar approach was used to characterize PEA-derived fragments exocytotically released into the extracellular space that showed five additional minor PEAPs. Among all these naturally generated peptides, enkelytin, the antibacterial bisphos- phorylated C-terminal peptide (PEA209-237), was predominantly generated, as shown by MALDI-TOF mass spectrometry analysis, which constituted an efficient method for its identification. Finally, the data on PEA intragranular and extracellular processing in adrenal medulla are discussed in regard to the known enzymatic processing mechanisms. We note the high conservation of the cleavage points in evolutionarily diverse organisms, highlighting an important biological function for the released PEAPs.

Phosphorylation and O-glycosylation sites of human chromogranin A (CGA79-439) from urine of patients with carcinoid tumors.

Because of their water-soluble properties, chromogranins (CGs) and chromogranin-derived fragments are released together with catecholamines from adrenal chromaffin cells during stress situations and can be detected in the blood by radiochemical and enzyme assays. It is well known that chromogranins can serve as immunocytochemical markers for neuroendocrine tissues and as a diagnostic tool for neuroendocrine tumors. In 1993, large CGA-derived fragments have been shown to be excreted into the urine in patients with carcinoid tumors and the present study deals with the characterization of the post-translational modifications (phosphorylation and O-glycosylation) located along the largest natural CGA-derived fragment CGA79-439. Using mild proteolysis of peptidic material, high performance liquid chromatography, sequencing, and mass spectrometry analysis, six post-translational modifications were detected along the C-terminal CGA-derived fragment CGA79-439. Three O-linked glycosylation sites were located in the core of the protein on Thr163, Thr165, and Thr233, consisting in di-, tri-, and tetrasaccharides. Three phosphorylation sites were located in the middle and C-terminal domain, on serine residues Ser200, Ser252, and Ser315. These modified sites were compared with sequences of others species and discussed in relation with the post-translational modifications that we have reported previously for bovine CGA.

[Blood cadmium and plasma zinc in hypertensive patients. Apropos of 76 cases]. / Cadmium sanguin et zinc plasmatique chez l'hypertendu. A propos de 76 observations.

Changes in total blood cadmium level (CdT), in blood plasma Zinc level (ZnP) and in CdT/ZnP ratio were studied in 39 hypertensive patients compared to 37 healthy subjects. The hypertensive patients were caracterized by a higher CdT level (p less than 10(-5)), a lower ZnP level (p less than 10(-7)) and a higher CdT/ZnP ratio (p less than 10(-7)). A negative correlation was found between CdT and ZnP, and between blood pressure values (systolic and diastolic) and ZnP. A positive correlation was found between blood pressure values and CdT, and therefore CdT/ZnP ratio.

[Carbonic anhydrase activity in male chronic alcoholics. Statistical analysis of correlations apropos of 74 cases]. / Activité de l'anhydrase carbonique chez l'éthylique chronique de sexe masculin. Analyse statistique des corrélations à propos de soixante-quatorze observations.

Changes in cadmium blood levels and erythrocyte contents and in carbonic anhydrase activity (C.A.A.) were studied comparatively in 74 male subjects: 37 chronic alcoholics and 37 healthy age-matched controls. Chronic alcoholics exhibited significantly higher serum and erythrocyte cadmium concentrations and lower CAA than controls. Cadmium levels were correlated with the CAA. There was a negative correlation between the CAA and the mean corpuscular volume, daily alcohol intake and duration of the intemperance, in contrast with a positive correlation demonstrated for cadmium concentrations.