RESUMEN
Human papillomavirus (HPV) infection is the most common sexually transmitted disease in the world and is related to the etiology of cervical cancer. The most common high-risk HPV types are 16 and 18; however, the second most prevalent type in the Midwestern region of Brazil is HPV-33. New vaccine strategies against HPV have shown that virus-like particles (VLP) of the major capsid protein (L1) induce efficient production of antibodies, which confer protection against the same viral type. The methylotrophic yeast Pichia pastoris is an efficient and inexpensive expression system for the production of high levels of heterologous proteins stably using a wild-type gene in combination with an integrative vector. It was recently demonstrated that P. pastoris can produce the HPV-16 L1 protein by using an episomal vector associated with the optimized L1 gene. However, the use of an episomal vector is not appropriate for protein production on an industrial scale. In the present study, the vectors were integrated into the Pichia genome and the results were positive for L1 gene transcription and protein production, both intracellularly and in the extracellular environment. Despite the great potential for expression by the P. pastoris system, our results suggest a low yield of L1 recombinant protein, which, however, does not make this system unworkable. The achievement of stable clones containing the expression cassettes integrated in the genome may permit optimizations that could enable the establishment of a platform for the production of VLP-based vaccines.
Asunto(s)
Alphapapillomavirus/inmunología , Proteínas de la Cápside/biosíntesis , Proteínas Oncogénicas Virales/biosíntesis , Pichia/metabolismo , Alphapapillomavirus/genética , Anticuerpos Antivirales/inmunología , Proteínas de la Cápside/genética , Transformación Celular Viral/fisiología , Electroforesis en Gel de Poliacrilamida , Regulación Viral de la Expresión Génica , Proteínas Oncogénicas Virales/genética , Vacunas contra Papillomavirus/inmunología , Pichia/genética , Pichia/virología , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
O trabalho objetivou avaliar quatro níveis de restrição alimentar (RA) (0, 5, 10 e 15 por cento) e dois níveis de energia líquida (EL) na dieta (2.083 e 2.252kcal EL/kg de ração) sobre as características de carcaça de suínos na fase de terminação. Foram utilizados 48 suínos Dalland, 24 machos castrados e 24 fêmeas, distribuídos em delineamento experimental de blocos ao acaso em arranjo fatorial 4 2 2 (nível de restrição x nível de energia x sexo). Foram avaliados: peso da carcaça quente, comprimento da carcaça, espessura média do toucinho, rendimentos da carcaça e de cortes e relação carne:gordura na carcaça. Os resultados mostraram melhora na qualidade da carcaça, considerando-se a redução na espessura de toucinho (Y= 2,587 - 0,0186RA) e o aumento no peso (Y= 6,628 + 0,0276RA) e nos rendimentos do lombo (Y= 6,5895 + 0,1389RA - 0,0081RA²) e do pernil (Y= 17,143 + 0,0646RA) com o aumento da restrição alimentar. O aumento da energia líquida da dieta resultou em aumento no rendimento da carcaça (77,4 e 79,1 por cento). As fêmeas apresentaram menor espessura de toucinho do que os machos castrados (2,36 e 2,54 cm).