Green synthesis of silica-coated magnetic nanocarriers for simultaneous purification-immobilization of ß-1,3-xylanase.

Tailoring magnetic nanocarriers with tunable properties is of great significance for the development of multifunctional candidate materials in numerous fields. Herein, we report a one-pot biomimetic silicification-based method for the synthesis of silica-coated magnetic nanoparticles. The synthesis process was mild, low cost, and highly efficient, which took only about 21 min compared with 4.5-120 h in other literature. Then, the carriers had been characterized by VSM, SEM, TEM, XRD, FT-IR, and EDS to confirm their function. To evaluate the usefulness of the carriers, they were adopted to couple the purification and immobilization of ß-1,3-xylanase from the cell lysate in a single step with high immobilization yield (92.8 %) and high activity recovery (82.4 %). The immobilized enzyme also retained 58.4 % of the initial activity after 10 cycles and displayed good storage properties, and improved thermal stability, which would be promising in algae biomass bioconversion as well as other diverse applications.

A new approach for purification of the catalytic site of the angiotensin-conversion enzyme, N-domain, mediated by the ELP-Intein system.

Angiotensin-converting enzyme I (ACE) is a key part of the renin-angiotensin system. Its main function is to regulate blood pressure and the balance of salts in the body. Somatic ACE has two domains, N-C-, each of which has a catalytic site that exhibits 60%sequence identity. The N-domain has a specific action in the hydrolysis of beta-amyloid bodies and angiotensin (1-7), which activates the MAS receptor and triggers anti-thrombotic and anti-inflammatory actions. Our goal was to obtain the catalytic site Ala361 to Gly468 of the N domain region, csACEN, without needing purification by chromatography. We employed a method that uses an Elastin-like Polypeptide (ELP) and Intein sequences linked to the peptide of interest. The more differential for obtaining the pure peptide was the cultivation temperatures in the synthesis of ELPcsACEN at 37 °C, with a significant increase in expression. In the purification by ELP precipitation, we recorded the highest efficiency in the concentrations of 0.57 M and 0.8 M of ammonium sulfate buffer. Intein autocleavage study allows removal of the ELP sequence at acidic pH, with the buffers MES and Tris-HCl The present study defined the best conditions for obtaining pure csACEN that the literature has not yet described for peptides. Obtaining pure csACEN aims at future studies for therapeutic use in hypertension, Alzheimer's, and oncology.

Transient Expression of Dengue Virus NS1 Antigen in Nicotiana benthamiana for Use as a Diagnostic Antigen.

Dengue is a viral disease that represents a significant threat to global public health since billions of people are now at risk of infection by this mosquito-borne virus. The implementation of extensive screening tests is indispensable to control this disease, and the Dengue virus non-structural protein 1 (NS1) is a promising antigen for the serological diagnosis of dengue fever. Plant-based systems can be a safe and cost-effective alternative for the production of dengue virus antigens. In this work, two strategies to produce the dengue NS1 protein in Nicotiana benthamiana leaves were evaluated: Targeting NS1 to five different subcellular compartments to assess the best subcellular organelle for the expression and accumulation of NS1, and the addition of elastin-like polypeptide (ELP) or hydrophobin (HFBI) fusion tags to NS1. The transiently expressed proteins in N. benthamiana were quantified by Western blot analysis. The NS1 fused to ELP and targeted to the ER (NS1 ELP-ER) showed the highest yield (445 mg/kg), approximately a forty-fold increase in accumulation levels compared to the non-fused protein (NS1-ER), representing the first example of transient expression of DENV NS1 in plant. We also demonstrated that NS1 ELP-ER was successfully recognized by a monoclonal anti-dengue virus NS1 glycoprotein antibody, and by sera from dengue virus-infected patients. Interestingly, it was found that transient production of NS1-ER and NS1 ELP-ER using vacuum infiltration of whole plants, which is easier to scale up, rather than syringe infiltration of leaves, greatly improved the accumulation of NS1 proteins. The generated plant made NS1, even without extensive purification, showed potential to be used for the development of the NS1 diagnostic tests in resource-limited areas where dengue is endemic.