RESUMEN
Alginate-based microencapsulation of live cells may offer the opportunity to treat chronic and degenerative disorders. So far, a thorough assessment of physical-chemical behavior of alginate-based microbeads remains cloudy. A disputed issue is which divalent cation to choose for a high performing alginate gelling process. Having selected, in our system, high mannuronic (M) enriched alginates, we studied different gelling cations and their combinations to determine their eventual influence on physical-chemical properties of the final microcapsules preparation, in vitro and in vivo. We have shown that used of ultrapure alginate allows for high biocompatibility of the formed microcapsules, regardless of gelation agents, while use of different gelling cations is associated with corresponding variable effects on the capsules' basic architecture, as originally reported in this work. However, only the final application which the capsules are destined to will ultimately guide the selection of the ideal, specific gelling divalent cations, since in principle there are no capsules that are better than others.
Asunto(s)
Alginatos/química , Materiales Biocompatibles/uso terapéutico , Trasplante de Células/métodos , Composición de Medicamentos/métodos , Alginatos/uso terapéutico , Materiales Biocompatibles/química , Cápsulas/química , Cápsulas/uso terapéutico , Geles/química , Ácido Glucurónico/química , Ácido Glucurónico/uso terapéutico , Ácidos Hexurónicos/química , Ácidos Hexurónicos/uso terapéutico , HumanosRESUMEN
A new method for oxidative folding of synthetic polypeptides assembled by stepwise solid phase synthesis is introduced. Folding is obtained in excellent yields by reacting S-tert-butylthiolated polypeptides with a 100-fold molar excess of cysteine at 37 degrees C in a slightly alkaline buffer containing chaotropic salts, and in the presence of air-oxygen. This novel protocol has been applied to the folding of S-tert-butylthiolated human thymus and activation-regulated chemokine (hu-TARC) derivatives as well as to larger segments of Plasmodium falciparum and Plasmodium berghei circumsporozoite proteins. Folded P. falciparum polypeptides have been used as substrates of endoproteinase Glu-C (Glu-C) and endoproteinase Asp-N (Asp-N) in an attempt to identify their disulfide connectivities. Particular practical advantages of the present method are (i) easy purification and storage of the S-protected peptide derivatives, (ii) elimination of the risk of cysteine alkylation during the acidolytic cleavage deprotection and resin cleavage steps, (iii) possibility to precisely evaluate the extent of folding and disulfide bond formation by mass spectrometry, and (iv) facile recovery of the final folded product.
Asunto(s)
Péptidos/química , Pliegue de Proteína , Secuencia de Aminoácidos , Animales , Quimiocina CCL17/química , Cromatografía Líquida de Alta Presión , Humanos , Datos de Secuencia Molecular , Oxidación-Reducción , Péptidos/síntesis química , Péptidos/metabolismo , Plasmodium berghei/metabolismo , Plasmodium falciparum/metabolismo , Proteínas Protozoarias/química , Espectrometría de Masa por Láser de Matriz Asistida de Ionización DesorciónRESUMEN
Amphiphilic peptidyl-RNA conjugates, molecules that mimic natural peptidyl-transfer RNA, are capable of self-assembling on glass substrates as vesicles and supported bilayers.
Asunto(s)
Péptidos/química , ARN/química , Modelos Moleculares , Conformación de Ácido Nucleico , Conformación ProteicaRESUMEN
[reaction: see text] An efficient procedure for the immobilization of 3'-deoxy-3'-(O-methyltyrosyl)aminoadenosine was developed. A poly(ethylene glycol)-derived diacid linker/spacer was attached to aminomethyl polystyrene. Coupling of the 2'-hydroxy instead of the 2'-O-succinylated ribonucleoside resulted in high immobilization yields (over 80%) and allowed for the recovery of valuable unreacted material. This specific procedure should be applicable to other ribonucleosides containing a bulky modification at the 3'-position and can be used for the stepwise construction of 3'-aminoacyl- or 3'-peptidyl-RNA conjugates.
Asunto(s)
Puromicina/análogos & derivados , Aminoacil-ARN de Transferencia/síntesis química , Adenosina/análogos & derivados , Hidroxilación , Aminoacil-ARN de Transferencia/químicaRESUMEN
3'-aminoacylamino-3'-deoxyadenosines, analogues of the antibiotic puromycin, have been synthesized from adenosine. They key 3'-azido derivative 10 was obtained through a 3'-oxidation/reduction/substitution procedure. A modified purification protocol on a larger scale was developed for the oxidation step using the Garegg reagent. The coupling reaction between an Fmoc-l-amino acid and the fully protected form of 3'-amino-3'-deoxyadenosine 11 furnished the aminoacylated compounds 12 in high yields. The puromycin analogues were obtained in 10 steps and up to 23% (14c) overall yield.
Asunto(s)
Desoxiadenosinas/síntesis química , Puromicina/análogos & derivados , Puromicina/síntesis química , Adenosina , Química Orgánica/métodos , Desoxiadenosinas/análisis , Indicadores y Reactivos , Espectroscopía de Resonancia Magnética , Estructura Molecular , Puromicina/análisis , Estereoisomerismo , Relación Estructura-ActividadRESUMEN
The present work describes the recognition of three synthetic polypeptides encompassing the N- and C-terminal regions of the transmembrane Exp-1 protein of the parasite Plasmodium falciparum by plasma and peripheral blood mononuclear cells from naturally exposed individuals living in African endemic areas. The three polypeptides comprise the sequences 23-105, 73-162 and 101-162, and overlap at the transmembrane domain (73-105). Thus, they permitted characterization of the immune response specific to the N- and C-terminal domains in an independent fashion. Two different populations were evaluated, one in the village of Safo in Mali and the other in the villages of Somnaway, Kabortenga and Toussouktenga in Burkina Faso. Antibodies to the sequence 73-162 of Pf Exp-1 were found in 70% of adult Mali donors and in all of the donors tested from Burkina Faso. Strikingly, the N-terminal fragment Pf Exp-1 23-105 was only weakly recognized by a few donors. Evaluation of the T-cell response indicated that the peptide Pf Exp-1 23-105 was more potent than Pf Exp-1 73-162 in inducing a proliferative response. A correlation between peptide-specific interferon-gamma and interleukin-6 production and proliferation to peptide Pf Exp-1 23-105 was observed. Further studies are needed to evaluate this molecule as a vaccine candidate.