RESUMO
We demonstrate that a donut-shaped surface pattern consisting of a central hydrophobic region and a surrounding hydrophilic region simultaneously concentrates and desalts a solution of neuropeptides with a high salt content. Our approach greatly simplifies the sample preparation process for MALDI mass spectrometry.
Assuntos
Neuropeptídeos/química , Águas Salinas/química , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz/métodos , Angiotensina I/química , Bradicinina/análogos & derivados , Bradicinina/química , Peptídeo da Parte Intermédia da Adeno-Hipófise Semelhante à Corticotropina/química , Ouro/química , Interações Hidrofóbicas e Hidrofílicas , Cinética , Limite de Detecção , Modelos Químicos , Silício/química , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz/instrumentação , Compostos de Sulfidrila/químicaRESUMO
The remarkable conservation of the primary structures and anatomical location of dogfish α-melanocyte-stimulating hormone (MSH), corticotrophin-like intermediate lobe peptide (CLIP) and adrenocorticotrophic hormone (ACTH) compared with mammals reinforced the tissue-specific processing hypothesis of ACTH peptides in the pituitary gland. The cloning of dogfish pro-opiomelanocortin (POMC) led to the identification of δ-MSH and simultaneously revealed the high conservation of the γ-MSH sequence during evolution. These studies have also shown that ß-MSH is much less conserved during evolution and in some species is not even processed from ß-LPH. Human pro-γ-MSH potentiates the corticosteroidogenic activity of ACTH and peptides generated from its N-terminal, in particular big-γ-MSH, appear to have adrenal mitogenic activity. Human big-γ-MSH (from the zona intermedia) may also cause the adrenache. The review finishes with a cautionary note with regard to the misdiagnosis of the ectopic ACTH syndrome in which partial processing of ACTH can result in large concentrations of α-MSH and CLIP, which can interfere in the performance of two-site immunoassays, and the problem of the correct disulphide bridge arrangement in synthetic N-POMC peptides is also discussed.
Assuntos
Hormônio Adrenocorticotrópico/isolamento & purificação , Hormônios Estimuladores de Melanócitos/isolamento & purificação , Pró-Opiomelanocortina/isolamento & purificação , Síndrome de ACTH Ectópico/sangue , Síndrome de ACTH Ectópico/metabolismo , Glândulas Suprarrenais/metabolismo , Hormônio Adrenocorticotrópico/sangue , Hormônio Adrenocorticotrópico/química , Hormônio Adrenocorticotrópico/genética , Animais , Peptídeo da Parte Intermédia da Adeno-Hipófise Semelhante à Corticotropina/química , Peptídeo da Parte Intermédia da Adeno-Hipófise Semelhante à Corticotropina/genética , Peptídeo da Parte Intermédia da Adeno-Hipófise Semelhante à Corticotropina/isolamento & purificação , História do Século XX , Humanos , Hormônios Estimuladores de Melanócitos/sangue , Hormônios Estimuladores de Melanócitos/química , Hormônios Estimuladores de Melanócitos/genética , Hipersecreção Hipofisária de ACTH/sangue , Hipersecreção Hipofisária de ACTH/metabolismo , Hipófise/metabolismo , Pró-Opiomelanocortina/química , Pró-Opiomelanocortina/genética , Pró-Opiomelanocortina/história , Isoformas de Proteínas , alfa-MSH/química , alfa-MSH/genética , alfa-MSH/isolamento & purificação , beta-Endorfina/química , beta-Endorfina/genética , beta-Endorfina/isolamento & purificaçãoRESUMO
Drug/polymer interactions occur during in situ polymerization of poly(alkylcyanoacrylate) (PACA) formulations. We have used MALDI ionization coupled tandem time-of-flight (TOF) mass spectrometry as an accurate method to characterize covalent peptide/polymer interactions of PACA nanoparticles with the bioactives D-Lys6-GnRH, insulin, [Asn1-Val5]-angiotensin II, and fragments of insulin-like growth factor 1 (IGF-1 (1-3)) and human adrenocorticotropic hormone (h-ACTH, (18-39)) at the molecular level. Covalent interactions of peptide with alkylcyanoacrylate were identified for D-Lys6-GnRH, [Asn1-Val5]-angiotensin II and IGF-1 (1-3). D-Lys6-GnRH and [Asn1-Val5]-angiotensin II were modified at their histidine side chain within the peptide, whilst IGF-1 (1-3) was modified at the C-terminal glutamic acid residue. The more complex protein insulin was not modified despite the presence of 2 histidine residues. This might be explained by the engagement of histidine residues in the folding and sterical arrangement of insulin under polymerization conditions. As expected, h-ACTH (18-39) that does not contain histidine residues did not interfere in the polymerization process. Lowering the pH did not prevent the covalent association of PACA with D-Lys6-GnRH or IGF-1 (1-3). Conclusively, protein and peptide bioactives are potentially reactive towards alkylcyanoacrylate monomers via various mechanisms with limited interference of pH. Histidines and C-terminal glutamic acid residues have been identified as potential sites of interaction. The likelihood of their engagement in the polymerization process (initiators), however, seems dependent on their sterical availability. The reactivity of nucleophilic functional groups should always be considered and bioactives examined for their potential to covalently interfere with alkylcyanoacrylate monomers, especially when designing PACA delivery systems for protein and peptide biopharmaceuticals.
