Characterization of the oligosaccharide structure of native and recombinant human prolactinobtained from CHO cells treated or not with cycloheximide

• Grant number: 11/22985-3
• Support Opportunities: Regular Research Grants
• Start date: March 01, 2012
• End date: February 28, 2014
• Field of knowledge: Biological Sciences - Biochemistry - Molecular Biology
• Principal Investigator: Carlos Roberto Jorge Soares
• Grantee: Carlos Roberto Jorge Soares
• Host Institution: Instituto de Pesquisas Energéticas e Nucleares (IPEN). Secretaria de Desenvolvimento Econômico (São Paulo - Estado). São Paulo , SP, Brazil
• Associated researchers: Maria Teresa de Carvalho Pinto Ribela ; Miriam Fussae Suzuki ; Paolo Bartolini

Abstract

Human prolactin (hPRL) is a polypeptide hormone secreted by the anterior pituitary under the regulation of the hypothalamus and involved in a variety of biological processes, such as mammary gland development and lactation. The recombinant product is important for diagnosis and for the therapy of lactation insufficiency This hormone can be non-glycosylated (NG-hPRL) and glycosylated (G-hPRL), with molecular weight of approximately 23 and 25 kDa, respectively; it has a single N-glycosylation site (ASP 31), which is partially occupied, thus representing a particularly interesting model of glycosylation. The biological activity of G-hPRL is lower compared to non-glycosylated prolactin (~ 4 times), its physiological function is not well defined and the portion of carbohydrate appears to play an important role in the biosynthesis, secretion, biological activity and survival of the plasma hormone. In the presence of cicloheximide (CHX) we already obtained an increase of ~4 times in absolute value in the expression of G-hPRL, while the production of the non-glycosylated isoform was reduced. A study on the oligosacharidic composition of recombinant G-hPRL in comparison with the natural hormone, synthesized by the pituitary was, however, never carried out and never was studied the influence of CHX on the same carbohydrate structures. This will be the object of this study, starting from suspension adapted CHO cells, that provide on easy scale upgrading, an increase in cellular density and, consequently, of hormone production. The biological activity will be evaluated in vitro using BaF3/LLP cells, while pharmacokinetic studies will also be carried out. (AU)

Scientific publications

(References retrieved automatically from Web of Science and SciELO through information on FAPESP grants and their corresponding numbers as mentioned in the publications by the authors)

CAPONE, MARCOS V. N.; SUZUKI, MIRIAM F.; OLIVEIRA, JOAO E.; DAMIANI, RENATA; SOARES, CARLOS R. J.; BARTOLINI, PAOLO. N-glycoprofiling analysis in a simple glycoprotein model: A comparison between recombinant and pituitary glycosylated human prolactin. Journal of Biotechnology, v. 202, p. 78-87, MAY 20 2015. (11/22985-3)