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In vivo modification of the goat mammary gland glycosylation pathway.
Leiva-Carrasco, María J; Jiménez-Chávez, Silvana; Harvey, David J; Parra, Natalie C; Tavares, Kaio C; Camacho, Frank; González, Alain; Sánchez, Oliberto; Montesino, Raquel; Toledo, Jorge R.
Afiliação
  • Leiva-Carrasco MJ; Biotechnology and Biopharmaceuticals Laboratory, Pathophysiology Department, School of Biological Sciences, University of Concepcion, Victor Lamas 1290, P.O. Box 160C, Concepcion, Chile; Biotechnology and Biomedicine Center SpA, Granada 168, Villumanque, Concepcion, Chile.
  • Jiménez-Chávez S; Biotechnology and Biopharmaceuticals Laboratory, Pathophysiology Department, School of Biological Sciences, University of Concepcion, Victor Lamas 1290, P.O. Box 160C, Concepcion, Chile; Biotechnology and Biomedicine Center SpA, Granada 168, Villumanque, Concepcion, Chile.
  • Harvey DJ; Oxford Glycobiology Institute, Biochemistry Department, South Parks Road, Oxford, OX1 3QU, UK.
  • Parra NC; Department of Pharmacology, School of Biological Sciences, University of Concepcion, Victor Lamas 1290, P.O. Box 160C, Concepcion, Chile.
  • Tavares KC; Molecular and Developmental Biology Laboratory, Experimental Biology Center (NUBEX), University of Fortaleza (UNIFOR), Fortaleza, CE, Brazil.
  • Camacho F; Department of Pharmacology, School of Biological Sciences, University of Concepcion, Victor Lamas 1290, P.O. Box 160C, Concepcion, Chile.
  • González A; Biotechnology and Biopharmaceuticals Laboratory, Pathophysiology Department, School of Biological Sciences, University of Concepcion, Victor Lamas 1290, P.O. Box 160C, Concepcion, Chile.
  • Sánchez O; Department of Pharmacology, School of Biological Sciences, University of Concepcion, Victor Lamas 1290, P.O. Box 160C, Concepcion, Chile.
  • Montesino R; Biotechnology and Biopharmaceuticals Laboratory, Pathophysiology Department, School of Biological Sciences, University of Concepcion, Victor Lamas 1290, P.O. Box 160C, Concepcion, Chile; Biotechnology and Biomedicine Center SpA, Granada 168, Villumanque, Concepcion, Chile. Electronic address: rmonte
  • Toledo JR; Biotechnology and Biopharmaceuticals Laboratory, Pathophysiology Department, School of Biological Sciences, University of Concepcion, Victor Lamas 1290, P.O. Box 160C, Concepcion, Chile; Biotechnology and Biomedicine Center SpA, Granada 168, Villumanque, Concepcion, Chile. Electronic address: jotole
N Biotechnol ; 61: 11-21, 2021 Mar 25.
Article em En | MEDLINE | ID: mdl-33157282
Complex recombinant glycoproteins produced as potential biopharmaceuticals in goat's milk have an aberrant pattern of N-glycosylation due to the lack of multi-antennary structures. Overexpression of glycosyltransferases may increase oligosaccharide branching of the desired glycoproteins. Here, human erythropoietin fused to human IgG Fc (EPO-Fc) was co-expressed with N-acetyl-glucosaminyltransferase-IVa (GnT-IVa) by adenoviral transduction in goat mammary gland to evaluate the in vivo modification of N-glycosylation pattern in this tissue. Adenoviral vectors, containing the EPO-Fc and GnT-IVa sequences were assembled for in vitro and in vivo expression in mammalian cell culture or in goat mammary gland. Protein detection was assessed by gel electrophoresis and western blot, and N-glycans were identified by HPLC and mass spectrometry. GnT-IVa overexpression and its colocalization with EPO-Fc in the Golgi apparatus of SiHa cells were demonstrated. N-glycan analysis of in vitro and in vivo expression of EPO-Fc modified by GnT-IVa (EPO-Fc/GnT-IVa) showed an increase in high molecular weight structures, which corresponded to tri- and tetra-antennary N-glycans in SiHa cells and mostly tri-antennary N-glycans in goat's milk from transformed mammary tissue. The results confirmed that successful modification of the goat mammary gland secretion pathway could be achieved by co-expressing glycoenzymes together with the glycoprotein of interest. This is the first report of modification of the N-glycosylation pattern in the goat mammary gland in vivo, and constitutes a step forward for improving the use of the mammary gland as a bioreactor for the production of complex recombinant proteins.
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Texto completo: 1 Coleções: 01-internacional Base de dados: MEDLINE Assunto principal: Glicoproteínas / Glândulas Mamárias Animais Limite: Animals / Female / Humans Idioma: En Revista: N Biotechnol Assunto da revista: BIOLOGIA MOLECULAR / ENGENHARIA BIOMEDICA Ano de publicação: 2021 Tipo de documento: Article País de afiliação: Chile País de publicação: Holanda
Texto completo
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Texto completo: 1 Coleções: 01-internacional Base de dados: MEDLINE Assunto principal: Glicoproteínas / Glândulas Mamárias Animais Limite: Animals / Female / Humans Idioma: En Revista: N Biotechnol Assunto da revista: BIOLOGIA MOLECULAR / ENGENHARIA BIOMEDICA Ano de publicação: 2021 Tipo de documento: Article País de afiliação: Chile País de publicação: Holanda