Expression, purification and thermal stability evaluation of an engineered amaranth protein expressed in Escherichia coli

Background: The acidic subunit of amarantin (AAC)-the predominant amaranth seed storage protein-has functional potential and its third variable region (VR) has been modified with antihypertensive peptides to improve this potential. Here, we modified the C-terminal in the fourth VR of AAC by inserting four VY antihypertensive peptides. This modified protein (AACM.4) was expressed in Escherichia coli. In addition, we also recombinantly expressed other derivatives of the amarantin protein. These include: unmodified amarantin acidic subunit (AAC); amarantin acidic subunit modified at the third VR with four VY peptides (AACM.3); and amarantin acidic subunit doubly modified, in the third VR with four VY peptides and in the fourth VR with the RIPP peptide (AACM.3.4). Results: E. coli BL21-CodonPlus (DE3)-RIL was the most favorable strain for the expression of proteins. After 6 h of induction, it showed the best recombinant protein titer. The AAC and AACM.4 were obtained at higher titers (0.56 g/L) while proteins modified in the third VR showed lower titers: 0.44 g/L and 0.33 g/L for AACM.3 and AACM.3.4, respectively. As these AAC variants were mostly expressed in an insoluble form, we applied a refolding protocol. This made it possible to obtain all proteins in soluble form. Modification of the VR 4 improves the thermal stability of amarantin acidic subunit; AAC manifested melting temperature (Tm) at 34°C and AACM.4 at 37.2°C. The AACM.3 and AACM.3.4 did not show transition curves. Conclusions: Modifications to the third VR affect the thermal stability of amarantin acidic subunit.

Chemical constituents of monoterpenes in fruits of Gardenia jasminoides / 中草药

Objective: To investigate the chemical constituents of monoterpenes in the fruits of Gardenia jasminoides. Methods: Various column chromatographies were used in the isolation and purification, and the physicochemical constant determination and spectral analysis were adopted to identify the chemical structures of monoterpenes. Results: Twelve monoterpenes were isolated from G. jasminoides, such as jasminoside B (1), jasminoside G (2), jasminodiol (3), crocusatin-C (4), (7S)-6-(hydroxymethyl)-1, 1, 5-trimethylcyclohex-3-enone (5), bornyl-6-O-β-D-xylopyranosyl-β-D-glucopyranoside (6), (10R, 11R)-gardendiol (7), (10S, 11S)-gardendiol (8), (5S, 9S)-gardenate A (9), (5R, 9R)-gardenate A (10), jasminoside E (11), and 5, 6-dihydroxymethyl-1, 1-dimethylcyclohex-4-enone (12). Conclusion: Compounds 5, 6, 8, 10, and 12 are first isolated from this plant.

Episodios clínicos de inquietud y pánico en los ataques terroristas del 11 de septiembre de 2001 en Estados Unidos / Clinical episodes of restlessness and panic in the terrorist attacks of 11 of September of 2001 in United States

Introducción: Conviene especificar que los nervios son una taxonomía biomédica antigua de la enfermedad, cuyas nociones se han reelaborado en contextos populares. Hoy es una enfermedad presente en muchas sociedades occidentales que expresan una manera del sufrimiento. Objetivo: Este estudio se centra en explorar el impacto del 11-S en los nervios de los inmigrantes mexicanos que viven en los Estados Unidos. Metodología: Está dirigida por la tradición de la antropología de la enfermedad. Se apoya en los materiales originales, recolectados en Highland Park (Illinois, Estados Unidos). El trabajo en el terreno se hizo de septiembre de 2001 a febrero de 2002 entre usuarios del "Programa Latino" de la Agencia "Servicio a la Familia de South Lake County". Los datos provienen de entrevistas recogidas en profundidad a los informadores calificados. Se consideraron tres dimensiones principales en la metodología y en el plan de trabajo. Primera, el impacto del episodio de las torres en la vida de cada día de los inmigrantes mexicanos, la manera en que su salud se afectó, y cómo se originaron y manifestaron los nervios. En segundo lugar, se consideran de forma particular algunas cuestiones como la incertidumbre sobre su futuro y su miedo, ligados no solamente a las posibilidades de nuevos ataques sino también en relación con los cambios internos y las políticas federales. En un tercer momento, se exploró cómo la identidad mexicana fue redefinida como consecuencia del ataque. Resultados: Ofrecen el dato que el padecimiento nervioso entre mexicanos inmigrantes en relación con la violencia política básicamente no difiere de las manifestaciones y teorías etiológicas señaladas por numerosos autores en relación con diversos grupos sociales, culturales y étnicos en trabajos previos.

Introduction: It agrees to specify that the nerves are an old biomedical taxonomy of the disease, whose slight knowledge has been elaborated in popular contexts. Today, it is a present disease in many western societies that express a way of the suffering. Objective: This study is centered in exploring the impact of 11-S in the nerves of the Mexican immigrants who live in the United States. Methodology: It is directed by the tradition of the anthropology of the disease. It leans in the original materials, collected in the park of the mountain (Illinois, the United States). The field work was lead from September of 2001 to February of 2002 between users of the Latin Program of the Agency service of the family of South Lake County. The data come from interviews gathered in depth to qualified informers. Three main dimensions were considered in the methodology and work plan. First, episode of the towers’s impact in every day life of Mexican immigrants, the way in which their health was affected, and how the nerves were originated and declared. Secondly, it considers of particular form some questions like the uncertainty on their future and the fear, bound not only to the possibilities of new attacks but also related to the internal changes and the federal policies. At a third moment, it was explored how the Mexican identity was redefined as a result of the attack. Results: They offer the data that the nervous suffering between Mexican immigrants regarding political violence basically does not defer from the manifestations and etiological theories indicated by numerous authors in relation to diverse social groups, cultural and ethnic in previous works.

Effect of alkaline pH on sunflower 11S protein.

The effect of alkaline pH on sunflower 11S protein has been monitored by the techniques of ultracentrifugation, polyacrylamide gel electrophoresis, turbidity, viscosity, ultraviolet absorption spectra and fluorescence spectra· Both ultracentrifugation and polyacrylamide gel electrophoresis show the dissociation of the protein with increase in pH. Turbidity values decrease with pH while viscosity increases· With increase in pH absorbance of the protein solution increases and there is a red shift in the absorption maximum. Fluorescence quenching and a red shift in the emission maximum are also observed. Both dissociation and denaturation of the protein occur. Analysis of turbidity, viscosity and fluorescence data suggests that apparently denaturation follows dissociation.

Transforming growth factor-? conjugated with cytotoxin saporin inhibits specifically proliferating vascular smooth muscle cells / 中国药理学通报

AIM To testify the special cytotoxicity of TGF? SAP on proliferating vascular smooth muscle cells and endothelial cells. METHODS Conjugation of saporin to TGF? was accomplished after derivatization of saporin and TGF? with N succinimidyl 3 (2 pyridyldithio) proprionate. Cytotoxicity assays were measured by cell count. The studies of influence of TGF? SAP on values of thymidine and leucine incorporation into SMCs and ECs were measured by 3H thymidine uptake and 3H leucine uptake, respectively. RESULTS Cytotoxicity assays testified TGF? SAP conjugate could inhibit remarkably proliferation of SMCs in culture. The values of thymidine of TGF? SAP group (1?10 -9 mol?L -1 and 1?10 -7 mol?L -1 ) in comparison significantly decreased to 60 9% and 56 0% of the control group respectively, suggesting that cellular DNA synthesis obviously decreased as TGF? SAP was added. But saporin did not affect cellular DNA synthesis at higher level. The rate of 3H leucine incorporation of TGF? SAP group significantly decreased to 47 3% of the control group, suggesting that SMCs protein synthesis obviously decreased as TGF? SAP was added. But TGF? SAP at the same level did not affect DNA synthesis and protein synthesis of ECs compared with the control group. CONCLUSION TGF? SAP possesses the more effective cytotoxicity than saporin and the more specific citotoxicity on proliferating vascular smooth muscle cells than on proliferating endothelial cells.

Effect of denaturants on the 11S protein of guar seed.

The effect of sodium dodecyl sulphate, urea or guanidinium hydrochloride on the sedimentation velocity, viscosity, ultra violet spectra and fluorescence spectra of the 11S protein of guar seed has been determined. Sodium dodecyl sulphate dissociates the protein directly to the 2S protein, whereas urea or guanidinium hydrochloride produces an intermediate 7S protein. These reagents denature the protein also. Both the dissociative and the denaturation effect follow the order, sodium dodecyl sulphate > guanidinium hydrochloride > urea when the concentration are expressed as mols per litre. The denatured states in the three cases probably differ.

Effect of denaturants on the oligomeric structure of the 11 S protein of sunflower (Helianthus annum).

The effect of sodium dodecyl sulphate, urea, guanidinium hydrochloride and heat on the oligomeric structure of the 11 S protein of sunflower has been determined. Sodium dodecyl sulphate directly dissociates the protein to 2 S subunits, whereas urea and guanidinium hydrochloride dissociate it through an intermediate 7 S protein. Heating the protein at 90°C for 20 min caused dissociation of the 11 S protein, without any precipitation.