RESUMO
Small single-chain variable fragments (scFv) are promising biomolecules to inhibit and neutralize toxins and to act as antivenoms. In this work, we aimed to produce a functional scFv-6009FV in the yeast Pichia pastoris, which inhibits the pure Cn2 neurotoxin and the whole venom of Centruroides noxius. We were able to achieve yields of up to 31.6 ± 2 mg/L in flasks. Furthermore, the protein showed a structure of 6.1 % α-helix, 49.1 % ß-sheet, and 44.8 % of random coil by CD. Mass spectrometry confirmed the amino acid sequence and showed no glycosylation profile for this molecule. Purified scFv-6009FV allowed us to develop anti-scFvs in rabbits, which were then used in affinity columns to purify other scFvs. Determination of its half-maximal inhibitory concentration value (IC50) was 40 % better than the scFvs produced by E. coli as a control. Finally, we found that scFv-6009FV was able to inhibit ex vivo the pure Cn2 toxin and the whole venom from C. noxius in murine rescue experiments. These results demonstrated that under the conditions assayed here, P. pastoris is suited to produce scFv-6009FV that, compared to scFvs produced by E. coli, maintains the characteristics of an antibody and neutralizes the Cn2 toxin more effectively.
Assuntos
Anticorpos de Cadeia Única , Animais , Camundongos , Coelhos , Sequência de Aminoácidos , Animais Peçonhentos , Anticorpos Neutralizantes/química , Anticorpos Neutralizantes/genética , Anticorpos Neutralizantes/isolamento & purificação , Anticorpos Neutralizantes/farmacologia , Expressão Gênica , Neurotoxinas/antagonistas & inibidores , Neurotoxinas/química , Neurotoxinas/genética , Pichia/genética , Pichia/metabolismo , Proteínas Recombinantes/genética , Saccharomycetales/genética , Saccharomycetales/metabolismo , Venenos de Escorpião/antagonistas & inibidores , Venenos de Escorpião/química , Venenos de Escorpião/genética , Escorpiões , Anticorpos de Cadeia Única/química , Anticorpos de Cadeia Única/genética , Anticorpos de Cadeia Única/isolamento & purificação , Anticorpos de Cadeia Única/farmacologiaRESUMO
The valence and conduction band offsets for both polar and nonpolar CuGaS2/CuAlSe2 and CuGaS2/ZnSe interfaces were studied here by the state-of-the-art first-principles calculations. Using the hybrid functional calculations, we show that the CuGaS2/CuAlSe2 and CuGaS2/ZnSe heterostructures in all interfaces form type II band alignment. The difference of valence and conduction band offsets is mainly due to lattice mismatch, generating stress in the interface and affecting the electronic properties of each material; meanwhile, the polarity configuration does not play an important role in these values. From the local density of states and the charge density, we can determine how the nature of the band alignments changes when the semiconductor conforms to each interface. This allows us to localize the electrons and holes at different sites of the interface.
RESUMO
We report a case of a pregnant patient with a twin of 29 gestational weeks, and partial hydatidiform mole with coexistent living fetus. She was admitted at gyneco-obstetric urgencies due to a mild preeclampsia that evolves to severe. She has a complicated vaginal delivery with obstetric bleeding and there was practiced a total abdominal hysterectomy. Newborn dies 30 days later due to secondary complications (extreme prematurity). Patient evolution was satisfactory and she is currently under follow-up therapy to gestational trophoblastic disease without clinical or biochemical evidence of persistence or recurrence.