Improvement of recombinant protein production by an anti-apoptotic protein from hemolymph of Lonomia obliqua

Vieira, Helena L.A.; Pereira, Ana Cláudia; Peixoto, Cristina; Moraes, Roberto Henrique Pinto; Alves, Paula M.; Mendonça, Ronaldo Zucatelli

doi:10.1007/s10616-010-9305-x

Cited by 11 publications

(12 citation statements)

References 17 publications

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“…Several proteins are also targets of the biotechnology industry due to their large commercial interest. In this context, the caterpillar Lonomia obliqua gained great prominence in biotechnology in Brazil, owing to the active properties identified in its venom and in its hemolymph (Veiga et al, 2005), which can interfere in blood coagulation and fibrinolysis (Veiga et al, 2003), enhance cell growth (Maranga et al, 2003), act as anti-apoptotic agent (Souza et al, 2005) improve recombinant protein production (Mendonca et al, , 2008Vieira et al, 2010) and demonstrate antiviral effect .…”

Section: Introductionmentioning

confidence: 99%

Expression of an antiviral protein from Lonomia obliqua hemolymph in baculovirus/insect cell system

et al. 2012

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The control of viral infections, mainly those caused by influenza viruses, is of great interest in Public Health. Several studies have shown the presence of active properties in the hemolymph of arthropods, some of which are of interest for the development of new pharmacological drugs. Recently, we have demonstrated the existence of a potent antiviral property in the hemolymph of Lonomia obliqua caterpillars. The aim of this study was to produce an antiviral protein in a baculovirus/Sf9 cell system. The resulting bacmid contains the sequence coding for the antiviral protein previously described by our group. Total RNA from L. obliqua caterpillars was extracted with Trizol and used in the reverse transcription assay with oligo(d)T primer followed by polymerase chain reactions (RT-PCR) with specific primers for the cDNA coding for the antiviral protein, based on the sequence deposited in the GenBank database. Restriction sites were inserted in the cDNA for ligation in the donor plasmid pFastBac1™. The recombinant plasmid was selected in Escherichia coli DH5α and subsequently used in the transformation of E. coli DH10Bac for the construction of the recombinant bacmid. This bacmid was used for the expression of the antiviral protein in the baculovirus/Sf9 cell system. After identifying the protein by western blot, activity tests were performed, showing that the purified recombinant protein was able to significantly reduce viral replication (about 4 logs). Studies on the optimization of the expression system for the production of this antiviral protein in insect cells are in progress.

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Section: Introductionmentioning

confidence: 99%

Expression of an antiviral protein from Lonomia obliqua hemolymph in baculovirus/insect cell system

et al. 2012

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“…These anti-VDAC antibodies also inhibit apoptosis induced by chemical agents, inducers of apoptosis such as etoposide, paclitaxel and staurosporine. However, Cartier et al (1994) We have reported earlier the presence of anti-apoptotic protein present in the hemolymph of Lonomia oblique with his supplementation in cell's culture inducing high levels of cell growth (Mendonça et al 2008;Souza et al 2005;Mendonça et al 2002) and that the cells viability can be maintained for longer periods (Vieira et al 2010;Souza et al 2005;Maranga et al 2003). However, contrary that occurs with the anti-apoptotic Bcl-2 family proteins (Shimizu et al 1999), has the ability to inhibit apoptosis in different types of cells, whether mammal, either insect, by different inducers (Souza et al 2005;Vieira et al 2010).…”

Section: Discussionmentioning

confidence: 99%

“…However, Cartier et al (1994) We have reported earlier the presence of anti-apoptotic protein present in the hemolymph of Lonomia oblique with his supplementation in cell's culture inducing high levels of cell growth (Mendonça et al 2008;Souza et al 2005;Mendonça et al 2002) and that the cells viability can be maintained for longer periods (Vieira et al 2010;Souza et al 2005;Maranga et al 2003). However, contrary that occurs with the anti-apoptotic Bcl-2 family proteins (Shimizu et al 1999), has the ability to inhibit apoptosis in different types of cells, whether mammal, either insect, by different inducers (Souza et al 2005;Vieira et al 2010). The present data suggests that the site of action of this protein could be similar to Bcl (x) proteins, since that, in all experiments, as we observed a blockade of apoptosis, with clear protection of mitochondrial membrane potential.…”

Section: Discussionmentioning

confidence: 99%

“The anti-apoptotic effect ofLonomia obliquahemolymph is associated with the mitochondria pathway”

Mendonça¹,

Martins²

2022

Preprint

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The apoptosis death is a very important factors in production processes that limited the industrial production of some proteins of economic interest. However, one of the forms to increase the cellular productivity would be inhibit or attenuate the cellular death. Recently we have demonstrated the presence of a potent anti-apoptotic protein in Lonomia obliqua hemolymph which extends the cell culture viability through apoptosis prevention. By the other side, has been reported that mitochondria have one important action in the apoptosis control process, being that mitochondria membrane permeabilization (MMP) can be an important stage in this process. MMP associated or not with the loss of the electrochemical potential of the mitochondria and alteration of the matrix is responsible for the inter membranes protein release (e.g. cytochrome c, the AIF, etc) of cytosol. The result obtained showed that the addition of a protein from Lonomia obliqua hemolymph in the culture lead to a prolongation of the cellular life (3-4 days) and the cells leading a high electrochemical potential of the mitochondria. This protein can has its action in mitochondria membrane, avoiding the loss of the membrane permeability and the Cytochrome-C release. As positive control, apoptosis death in these cultures was induced by 50 μm of t-BHP or 600 μm of H202. The presence of apoptosis was characterized by flow citometry, microscopy electronic and agarose gel electrophoresis. The potential electrochemical of the mitochondria was determined by JC-1, Hoechst 33324 and DIOC6. Cytochrome C was identified in cytosol by an anti-cytochrome antibody.

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“…Molecules that interfere in the apoptotic process may be used in the biotechnology industry, the control of cell death occurring in high density cultures performed in bioreactors is an important factor in production processes. Has been shown that hemolymph of the Lonomia obliqua ( Lepdoptera:Saturniidade ) is capable of inhibiting cell death in different models [ 1 - 3 ]. The objective of this study, is to identify the potential anti-apoptotic of a protein isolated from hemolymph of larvae of Megalopyge albicolis ( Lepidoptera: Megalopygidae ).…”

mentioning

confidence: 99%

Study of antiapoptotic effect of a protein isolated from Megalopyge albicolis (Lepidoptera: Megalopygidae) hemolymph

et al. 2014

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Improvement of recombinant protein production by an anti-apoptotic protein from hemolymph of Lonomia obliqua

Cited by 11 publications

References 17 publications

Expression of an antiviral protein from Lonomia obliqua hemolymph in baculovirus/insect cell system

Expression of an antiviral protein from Lonomia obliqua hemolymph in baculovirus/insect cell system

“The anti-apoptotic effect ofLonomia obliquahemolymph is associated with the mitochondria pathway”

Study of antiapoptotic effect of a protein isolated from Megalopyge albicolis (Lepidoptera: Megalopygidae) hemolymph

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