Characterization of multiple forms of porcine anterior pituitary pro-opiomelanocortin N-terminal glycopeptide (original) (raw)
Related papers
Proceedings of the National Academy of Sciences of the United States of America, 1981
A glycopeptide isolated in relatively large amounts from human pituitary glands was completely purified, and its sequence was determined. The primary sequence represents the NH2-terminal 76 amino acid residues of pro-opiomelanocortin (POMC). This important secretory product of POMC was shown to possess an interesting aldosterone-stimulating activity on a human adrenal aldosteronoma. It is O-glycosylated at Thr-45 and N-glycosylated at Asn-65. Only one sequence variation with the human genomic DNA was found. Furthermore, comparison with the other preferred cleavage sites of human POMC reveals that the pair of basic residues Lys-Arg represents the major sites of enzymatic maturation of this precursor molecule. This predicts a highly specific type of enzyme involved in the maturation of POMC in the anterior lobe of the human pituitary.
The molecular characterisation of chicken pituitary N-terminal pro-opiomelanocortin (POMC)
Molecular and Cellular Endocrinology, 1998
Monoclonal antibodies (Mabs) specifically recognizing the chicken pituitary corticotropes were used to isolate a population of closely related peptides from crude chicken pituitary extracts. A homogeneous N-terminal sequence homologous to the extreme N-terminus of mammalian and amphibian pro-opiomelanocortin (POMC) was revealed. Further physicochemical analysis proved the existence of a series of C-terminally truncated peptides including 3 major molecular species corresponding to Ser1-Gly64, Ser1-Arg73 and Ser1-Gly105 respectively. The two latter molecules were shown to be N-glycosylated at position Asn67, with mass spectrometric data indicating a carbohydrate structure of the oligomannose 5 type, in addition to two more complex structures. No evidence was found in favour of O-glycosylation on Ser47. Degenerated PCR primers were deduced from the above protein sequence and from the known chicken adrenocorticotropic hormone (ACTH) sequence. The nucleotide sequence obtained by reversed transcription PCR (RT-PCR) completely confirmed the new amino acid sequence data including pro-gamma-MSH, the joining peptide and ACTH.
Biochemical and Biophysical Research Communications, 1981
The isolation and complete purification of a novel human pituitary glycspeptide (HPGP) from whole pituitaries is described. Amino acid composition predicts a glycopeptide rich in glucosamine. Complete sequence determination showed it to be a 39 residues with an oligosaccharide chain attached at Asn residue No. 6. It exhibits marked sequence homology to previousiy isolated pig posterior pituitary glycopeptide and to whole pituitary glyeopeptides isolated from ox, sheep and pig. Based on these results and the presence of such s peptide in posterior pituitary it is suggested that it could form part of the N-terminal glycopeptide extension of the precursor of neurophysin-arginine vasopressin.
Endocrinology, 1997
Two complementary DNAs encoding distinct forms of POMC have been characterized in the trout pituitary. One of the POMC variants (POMC-A) possesses a C-terminal extension of 25 amino acids, which has no equivalent in other POMCs described to date. This C-terminal peptide contains three pairs of basic amino acids, suggesting that it may be the precursor of multiple processed peptides. In addition, the presence of a C-terminal glycine residue suggests that some of the processing products may be ␣-amidated. To characterize the molecular forms of the peptides generated from the C-terminal domain of trout POMC-A, we have developed specific antibodies against the C-terminal pentapeptide YHFQG and its ␣-amidated derivative YHFQ-NH 2. Immunocytochemical labeling of pituitary sections with antibodies against YHFQ-NH 2 revealed the presence of numerous immunoreactive cells in the pars intermedia and the rostral pars distalis. In contrast, the antibodies against YHFQG produced only weak immunostaining. HPLC analysis combined with RIA detection revealed that extracts of the pars intermedia and pars distalis contain several peptides derived from the C-terminal extension of trout POMC-A, with the predominant molecular form exhibiting the same retention time as ALGERKYHFQ-NH 2. Tryptic digestion of this major form produced a peptide that coeluted with YHFQ-NH 2. These data indicate that the processing of the C-terminal extension of trout POMC-A generates several novel peptides including the decapeptide amide ALGERKYHFQ-NH 2. (Endocrinology 138: 128-137, 1997) Materials and Methods Animals A total of 130 adult rainbow trout, Oncorhynchus mykiss, of both sexes were used in the present study. The animals were obtained from a fish
Endocrinology, 1997
Two complementary DNAs encoding distinct forms of POMC have been characterized in the trout pituitary. One of the POMC variants (POMC-A) possesses a C-terminal extension of 25 amino acids, which has no equivalent in other POMCs described to date. This C-terminal peptide contains three pairs of basic amino acids, suggesting that it may be the precursor of multiple processed peptides. In addition, the presence of a C-terminal glycine residue suggests that some of the processing products may be ␣-amidated. To characterize the molecular forms of the peptides generated from the C-terminal domain of trout POMC-A, we have developed specific antibodies against the C-terminal pentapeptide YHFQG and its ␣-amidated derivative YHFQ-NH 2 . Immunocytochemical labeling of pituitary sections with antibodies against YHFQ-NH 2 revealed the presence of numerous immunoreactive cells in the pars intermedia and the rostral pars distalis. In contrast, the antibodies against YHFQG produced only weak immunostaining. HPLC analysis combined with RIA detection revealed that extracts of the pars intermedia and pars distalis contain several peptides derived from the C-terminal extension of trout POMC-A, with the predominant molecular form exhibiting the same retention time as ALGERKYHFQ-NH 2 . Tryptic digestion of this major form produced a peptide that coeluted with YHFQ-NH 2 . These data indicate that the processing of the C-terminal extension of trout POMC-A generates several novel peptides including the decapeptide amide ALGERKYHFQ-NH 2 . (Endocrinology 138: 128 -137, 1997) FIG. 4. Microphotographs showing the distribution of YHFQG-immunoreactive cells in the pars intermedia (A) and the rostral pars distalis (B). Preincubation of the antiserum with synthetic YHFQG (10 Ϫ5 M) resulted in complete extinction of the immunoreaction in the pars intermedia (C) and rostral pars distalis (D). Scale bars ϭ 10 m.
Mass spectrometric charting of bovine posterior/intermediate pituitary peptides
Proceedings of the National Academy of Sciences, 1989
The feasibility for charting neuropeptides in neuroendocrine tissues on the basis of the universal property and inherent specificity of their molecular weights was explored. As a model, a comprehensive MS analysis ofextractable peptides from bovine posterior/intermediate pituitary was performed. Two suitable MS techniques-namely, plasmadesorption time-of-flight and fast atom bombardment MSwere evaluated, and each method could identify more than 20 peptides, including N-terminally acetylated and C-terminally amidated species. In toto these peptides account for almost the entire lengths of propressophysin, prooxyphysin, and proopiomelanocortin. Some of the experimentally determined molecular weights did not match any known peptides. Three of these species were identified as acidic joining peptide(4-24)
Molecular and Cellular Endocrinology, 1994
The existence of heterogeneous molecular species of pro-opiomelanocortin (POMC) has been reported and it has been inferred that this explains the distinct release patterns of POMC-derived peptide hormones by the anterior and intermediate lobes of the pituitary gland. The aim of this study was to determine the nucleotide sequences of porcine pituitary anterior and intermediate lobar POMC from animals of the same strain. The POMC cDNAs were obtained using immunoscreening (anterior lobe) and the polymerase chain reaction (intermediate lobe), and their nucleotide sequences determined. Comparisons of the coding and the 5'-untranslated regions of the two POMCs demonstrated that their nucleotide sequences were identical and Northern blot analysis showed that both mRNAs were the same length. Therefore, the results of this study confirm that the same POMC transcript is present in both the anterior and intermediate pituitary lobes. The differences between the nucleotide and amino acid sequences of porcine POMC found hitherto may be attributable to strain differences. Comparisons of porcine and several vertebrate POMCs revealed highly conserved amino acid sequences in the regions corresponding to the peptide hormones, but the regions between them show considerable evolutionary divergence.
Journal of Neurochemistry, 1989
Coordinate secretion of two prohormoneJproneuropeptide processing enzymes [pro-opiomelanocortin converting enzyme (PCE) and an aminopeptidase B-like enzyme (APBE)] and a-melanotropin (a-MSH) from bovine intermediate lobe pituitary cells was studied. Stimulation of secretion with 8-bromo-cyclic AMP produced significant increases in levels of immunoreactive a-MSH, PCE, and APBE. Treatment of cells with the dopaminergic agonist 2-bromoa-ergocryptine resulted in significant decreases in secretion of a-MSH, PCE, and APBE. In neither case were there significant changes in levels of cytosolic lactic dehydrogenase or lysosomal P-glucuronidase in the medium. The secreted PCE activity was shown to process frog and mouse pro-opiomelanocortin primarily to 23,000-M, corticotropin (ACTH), 13,000-M, ACTH, P-lipotropin, a @-endorphin-like peptide, and &endorphin, products comparable to those synthesized by the mouse and frog intermediate lobe in situ. The secreted enzymatic activity had a pH optimum between 4.0 and 5.0, was strongly inhibited by pepstatin A, and had an inhibitor profile similar to the purified bovine intermediate lobe PCE. The secreted APBE activity cleaved ArgO-[Met]-enkephalin to [Met]-enkephalin and had a pH optimum and inhibitor profile similar to that previously reported for an activity from purified secretory vesicle fractions of bovine intermediate and neural lobes. The coordinate regulated secretion of a-MSH and enzyme activities (PCE and APBE) strongly indicates their colocalization in the same secretory vesicle compartment within the cell. The characteristics of the two enzymes secreted in the medium paralleled those seen in the tissue and further support their role in pro-opiomelanocortin processing in vivo. Key Words: a-Melanotropin-Pro-opiomelanocortin converting enzyme-Aminopeptidase B-like enzyme-Pro-opiomelanocortin-Intermediate lobe pituitary cells-Coordinate enzyme secretion. Castro M. G. et al. Regulated secretion of pro-opiomelanocortin converting enzyme and an aminopeptidase B-like enzyme from dispersed bovine intermediate lobe. pituitary cells.