Page 7 - PR 2014 2016 04 Biotechnology

Biotechnology | Progress Report 93

with The Agency for Agribusiness Technology its analogs in a research laboratory scale. In
of Piracicaba (SP) testing different commercial this period, the receptor antagonist G120R-
preparations of equine chorionic gonadotropin hGH was developed. A publication in “Protein
(eCG) concerning ovarian stimulation in cattle Expression and Purification” (Menezes ACSC et
(Alvarez RH et al., 2016). al., 2017) describes the synthesis and charac-
terization of G120R-hGH secreted into bacterial
Pirarucu (Araipama gigas) periplasm and obtained with a vector based

pituitary gonadotropins on a constitutive lambda PL promoter. This
antagonist can be useful for studies aiming
Araipama gigas (pirarucu) is a giant fish native at investigating the effects of a simultane-
to the Amazon River basin that can reach up ous inhibition of GH and prolactin signaling,
to 2 meters in length and in weigh up to 150 as a potential anti-tumoral or anti-diabetic
Kg. This species is in danger of disappearing compound. Studies related to glycosylated
because of the exploitation by the fishing in- hPRL (G-hPRL) were published in Journal of
dustry and increasing human presence. It is Biotechnology (Capone MN et al., 2015). The
largely used for food and extractivism and N-glycan structures present in native pituitary
commercial breeding is still incipient. In the G-hPRL were determined and compared with
last period (2011-2013), our research group those present in the recombinant hormone.
has isolated, from the pituitaries, and charac- To obtain recombinant G-hPRL, genetically
terized the cDNA of the α-subunit of A. gigas modified Chinese hamster ovary cells (CHO)
(ag) gonadotropins (FSH and LH), carrying out adapted to growth in suspension were treat-
also a phylogenetic analysis based on its amino ed with cycloheximide, thus increasing the
acid sequence. During this period (2014-2016), glycosylation site occupancy from 5.5% to
the cDNAs of ag-FSHβ and ag-LHβ-subunits 38.3%, thereby facilitating G-hPRL purifica-
have been isolated, characterized and phylo- tion. N-Glycan profiling proved to be a useful
genetically analysed. These results, including and accurate methodology also for monosac-
a three dimensional comparative modeling of charide determination, molecular mass and
ag-FSH and ag-LH, were submitted to publica- carbohydrate content determination for the
tion and are under revision by the Journal Plos two G-hPRL preparations, in good agreement
One. The biotechnological synthesis of these with the values obtained directly via MALDI-
hormones, useful for physiology and fertility TOF-MS.
applications, will be carried out with basis on
these isolated genes. Cellular response to

Prolactin (PRL) ionizing radiation

Ionizing radiation is a physical agent known
Prolactin is a 23 kDa protein hormone secreted to induce mutation and cancer, being also
by the anterior pituitary. Its therapeutic use is used as a widespread therapeutic modality for
still limited, while it is important for diagnostic cancer treatment. Thus, one of the challenges
applications such as prolactinomas, infertility in radiobiology and oncology is to understand
and tumorigenesis. In the context of recom- how the cells respond to radio-induced stress
binant hormone production, the group has because of exposure to radiation and its out-
been developing the expression, purification comes. The group, aiming to assess radio-in-
and characterization of human prolactin and duced cytogenetic damage/mutagenesis has