A comparison between the recombinant expression and chemical synthesis of a short cysteine-rich insecticidal spider peptide

Background:The choice between heterologous expression versus chemical synthesis for synthesizing short cysteine-rich insecticidal peptides from arthropods may impact the obtainment of yields and well-folded bioactive molecules for scientific research. Therefore, two recombinant expression systems were compared to that of chemical synthesis for producing Ba1, a cysteine-rich spider neurotoxin.Methods:The transcription of the insecticidal neurotoxin Ba1 was obtained from a cDNA library of venom glands of the spider Brachypelma albiceps.It was cloned into the pCR®2.1-TOPO® cloning vector and then introduced in two different expression vectors, pQE40 and pET28a+. Each vector was transfected into E. coli M15 and BL21 cells, respectively, and expressed under induction with isopropyl thiogalactoside (IPTG). The chemical synthesis of Ba1 was performed in an Applied Biosystems 433A peptide synthesizer.Results:Both expression systems pQE40 and pET28a+ expressed the His-tagged recombinant protein products, HisrDFHRBa1 and HisrBa1, respectively, as inclusion bodies. The recombinant proteins HisrDFHRBa1 and HisrBa1 presented respective molecular masses of 28,289 and 8274.6 Da, and were not biologically active. These results suggested that both HisrDFHRBa1 and HisrBa1 were oxidized after cell extraction, and that their insecticidal activities were affected by their N-terminal pro-peptides and different disulfide bridge arrangements. The respective protein expression yields for HisrDFHRBa1 and HisrBa1 were 100 μg/L and 900 μg/L of culture medium. HisrBa1 was reduced and folded under in vitroconditions. The in vitro folding of HisrBa1 produced several isoforms, one of which, after removing its N-terminal pro-peptide by enzymatic cleavage, presented elevated insecticidal activities compared to the native Ba1. Furthermore, the His-tagged protein HisrDFHRBa1 underwent enzymatic cleavage to obtain recombinant Ba1 (rBa1). As expected, the molecular mass of rBa1 was 4406.4 Da. On the other hand, Ba1 was chemically synthesized (sBa1) with a yield of 11 mg per 0.1 mmol of amino acid assembly.Conclusions:The two recombinant insecticidal peptides and the one synthesized chemically were as active as the native Ba1; however, toxin yields differed drastically.(AU)

A comparison between the recombinant expression and chemical synthesis of a short cysteine-rich insecticidal spider peptide

Background: The choice between heterologous expression versus chemical synthesis for synthesizing short cysteine-rich insecticidal peptides from arthropods may impact the obtainment of yields and well-folded bioactive molecules for scientific research. Therefore, two recombinant expression systems were compared to that of chemical synthesis for producing Ba1, a cysteine-rich spider neurotoxin. Methods: The transcription of the insecticidal neurotoxin Ba1 was obtained from a cDNA library of venom glands of the spider Brachypelma albiceps.It was cloned into the pCR®2.1-TOPO® cloning vector and then introduced in two different expression vectors, pQE40 and pET28a+. Each vector was transfected into E. coli M15 and BL21 cells, respectively, and expressed under induction with isopropyl thiogalactoside (IPTG). The chemical synthesis of Ba1 was performed in an Applied Biosystems 433A peptide synthesizer. Results: Both expression systems pQE40 and pET28a+ expressed the His-tagged recombinant protein products, HisrDFHRBa1 and HisrBa1, respectively, as inclusion bodies. The recombinant proteins HisrDFHRBa1 and HisrBa1 presented respective molecular masses of 28,289 and 8274.6 Da, and were not biologically active. These results suggested that both HisrDFHRBa1 and HisrBa1 were oxidized after cell extraction, and that their insecticidal activities were affected by their N-terminal pro-peptides and different disulfide bridge arrangements. The respective protein expression yields for HisrDFHRBa1 and HisrBa1 were 100 μg/L and 900 μg/L of culture medium. HisrBa1 was reduced and folded under in vitroconditions. The in vitro folding of HisrBa1 produced several isoforms, one of which, after removing its N-terminal pro-peptide by enzymatic cleavage, presented elevated insecticidal activities compared to the native Ba1. Furthermore, the His-tagged protein HisrDFHRBa1 underwent enzymatic cleavage to obtain recombinant Ba1 (rBa1). As expected, the molecular mass of rBa1 was 4406.4 Da. On the other hand, Ba1 was chemically synthesized (sBa1) with a yield of 11 mg per 0.1 mmol of amino acid assembly. Conclusions: The two recombinant insecticidal peptides and the one synthesized chemically were as active as the native Ba1; however, toxin yields differed drastically.

Massive hard coral loss after a severe bleaching event in 2010 at Los Roques, Venezuela / Pérdida masiva de coral duro después de un evento de blanqueamiento severo en 2010 en Los Roques, Venezuela

Thermal anomalies have become more severe, frequent and well-documented across the Caribbean for the past 30 years. This increase in temperature has caused coral bleaching resulting in reef decline. At Los Roques National Park, Venezuela, temperature has been monitored at four reef sites. In mid-September 2010, seawater temperature reached 30.85°C at 5 m depth in Los Roques, an archipelago only slightly affected by previous bleaching events. For example, bleaching in Los Roques in 2005 was mild compared to the rest of the Caribbean and to the results in this study. In 2010, seawater temperatures remained above 29.0°C from mid-August until the first week of November, resulting in +16 Degree Heating Weeks by that time. Our annual survey of four reef sites indicated that 72% of 563 scleractinian colonies were partial or totally bleached (white) or pale (discolored) in October 2010. In February 2011, there were still 46% of coral colonies affected; but most of them were pale and only 2% were bleached. By February, coral cover had declined 4 to 30% per transect, with a mean of 14.3%. Thus, mean coral cover dropped significantly from 45 to 31% cover (a 34% reduction). In addition to bleaching, corals showed a high prevalence (up to 16%) of black band disease in October 2010 and of white plague (11%) in February 2011. As a consequence, coral mortality is expected to be larger than reported here. Reef surveys since 2002 and personal observations for more than 20 years indicated that this bleaching event and its consequences in Los Roques have no precedent. Our results suggest that reef sites with no previous record of significant deterioration are more likely to become affected by thermal anomalies. However, this archipelago is relatively unaffected by local anthropogenic disturbance and has a high coral recruitment, which may contribute to its recovery.

Durante las últimas décadas las anomalías térmicas han sido más frecuentes y severas en el Caribe, quedando pocos arrecifes exentos de eventos masivos de blanqueamiento (EMB). En el Parque Nacional Los Roques, Venezuela, un archipiélago poco afectado previamente por EMB, la temperatura del agua a 5m de profundidad alcanzó 30,85°C en septiembre 2010, y fue >29,0°C entre mediados de agosto y la primera semana de noviembre en cuatro arrecifes. El 72% de 563 colonias de escleractinios estaban blanqueadas o pálidas para octubre de 2010, mientras que para febrero 2011, el 46% de las colonias aún estaban afectadas. Para febrero 2011, la cobertura béntica coralina promedio disminuyó de 45 a 31%. Además, los arrecifes mostraron una alta prevalencia (de hasta 16%) de enfermedad de banda negra en Octubre 2010, y de plaga blanca (11%) en Febrero 2011. Como consecuencia, es probable que la mortalidad coralina resulte mayor a la reportada acá. Sin embargo, Los Roques es poco afectado por perturbaciones antropogénicas y cuenta con un alto reclutamiento de corales, lo cual podría contribuir a su recuperación.

Quién es quién en las ciencias en Cuba: antecedentes y perspectivas / Who is wo in sciences in Cuba: background and perspectives

Se exponen los antecedentes, la evolución, así como las características esenciales del repertorio titulado ôQuién es quién en las ciencias en Cubaö, una obra de referencia, publicada en versión impresa y electrónica, que facilita la identificación de las más prominentes personalidades e instituciones de las ciencias cubanas, el intercambio de experiencias entre los especialistas y entidades, así como el desarrollo de estudios cienciológicos en diferentes disciplinas. Dirigido a los profesionales de la información, las instituciones de la ciencia, los medios de comunicación masiva, contiene los registros de unos 340 profesionales del sector de la salud y el medio ambiente. Es significativa la reducida cantidad de personalidades que aparecen registradas en el repertorio, si se considera la existencia de múltiples instituciones y personalidades en el país

Cuidados de enfermería en el recién nacido de bajo peso: factores asociados / Nursing care in the low-birth-weight newborn infants: associated factors

Se realiza el estudio de los 86 nacimientos ocurridos en el Hospital Maternoinfantil "Leopoldito Martínez" de San José de las Lajas" en el período comprendido entre el 1 de julio de 1982 y el 30 de junio de 1983. Se consideró como recién nacido de bajo peso lo propuesto por Batlagia y Lubchenco. Se analizaron un conjunto de factores que estuvieron presentes antes o durante el embarazo de los nacidos bajo peso. Se determinó el estado al nacer y las enfermedades más frecuentes. La mayor frecuencia de Apgar bajo ocurrió en los productos de embarazo con menos de 36 semanas. Las enfermedades más frecuentes en el bajo de peso fueron en orden descendente la hiperbilirrubinemia, la plétora sanguínea, el ictero fisiológico, la hipoglicemia, la sepsis urinaria, el distress respiratorio y la bronconeumonia congénita. Se concluye que es importante por parte del personal de Enfermería que labora en la Sala de neonatología aplicar los cuidados específicos a este recién nacido

Eliptocitosis herediataria no hemolítica en una familia mexicana / Herditary non-haemolytic elliptocytosis in a Mexican family

Se informa de hallazgos realizados en forma accidental en uno de los miembros de una familia mexicana con eliptocitosis hereditaria no hemolítica. De los 25 familiares estudiados, 15 presentaron la alteración. Esta enfermedad se diagnostica al observar en el frotis de sangre periférica más de 25% de eritrocitos con forma "eliptica". El diagnóstico se confirma cuando varios miembros de la familia están afectados

Anomalía de Relger-Huët: observaciones en una familia mexicana / Pelger Huët anomaly: observations in a Mexican family

Se informa de los hallazgos realizados en forma incidental en uno de los miembros de una familia mexicana con anomalía de Pelger-Huët. De los 29 familiares estudiados, 12 presentaron esta alteración. El diagnóstico de esta anomalía se establece en el estudio del frotis en sangre periférica y se confirma cuando varios miembros de una familia están afectados