Em 2009, foi demonstrado um potente antiviral na hemolinfa de L. obliqua. Esta proteína purificada reduziu o título viral (TCID50 ml-1) do sarampo, pólio e H1N1 em 157, 61 e 64 vezes. Recentemente, expressamos a proteína antiviral em sistema baculovírus, que reduziu o título de vírus do herpes em 106 vezes, da rubéola e EMC em 105 vezes. No entanto, este sistema de expressão é muito caro e trabalhoso. Com isso, clonamos e expressamos esta proteína com atividade antiviral em sistema bacteriano. A sequência antiviral foi clonada em vetor de expressão pET28a. As construções resultantes foram expressas em E. coli Bl21 DE3 pLysS induzidas com IPTG 1,0 mM. Após expressão, o pellet bacteriano foi sonicado e o material foi purificado por afinidade e gel filtração. Testou-se contra o vírus EMC mostrando uma proteção de 104 vezes em relação ao controle. E utilizando qPCR para determinar os níveis de transcrição de RNA do vírus do herpes e da rubéola em células infectadas mostrou uma inibição respectivamente de 106 e 105 vezes, em relação ao controle. / In 2009, we demonstrated a potent antiviral in the hemolymph of L. obliqua. This purified protein has reduced the viral titer (TCID50 ml-1), measles and polio H1N1 157, 61 and 64 times. Recently, the antiviral protein expressed in baculovirus system, which reduced the title of the herpes virus in 106 times, rubella and EMC in 105 times. However, this expression system is very expensive and laborious. Thus, cloned and expressed this protein with antiviral activity in bacterial system. The antiviral sequence was cloned into pET28a expression vector. The resulting constructs were expressed in E. coli BL21 DE3 pLysS cells induced with 1.0 mM IPTG. Following expression, the bacterial pellet was sonicated and the material was purified by affinity and gel filtration. Was tested against EMC virus showing a protection of 104 times compared to the control. And using qPCR to determine the levels of viral RNA transcription of the herpes virus and rubella-infected cells showed an inhibition of respectively 106 and 105 times compared to the control.

Em 2009, foi demonstrado um potente antiviral na hemolinfa de L. obliqua. Esta proteína purificada reduziu o título viral (TCID50 ml-1) do sarampo, pólio e H1N1 em 157, 61 e 64 vezes. Recentemente, expressamos a proteína antiviral em sistema baculovírus, que reduziu o título de vírus do herpes em 106 vezes, da rubéola e EMC em 105 vezes. No entanto, este sistema de expressão é muito caro e trabalhoso. Com isso, clonamos e expressamos esta proteína com atividade antiviral em sistema bacteriano. A sequência antiviral foi clonada em vetor de expressão pET28a. As construções resultantes foram expressas em E. coli Bl21 DE3 pLysS induzidas com IPTG 1,0 mM. Após expressão, o pellet bacteriano foi sonicado e o material foi purificado por afinidade e gel filtração. Testou-se contra o vírus EMC mostrando uma proteção de 104 vezes em relação ao controle. E utilizando qPCR para determinar os níveis de transcrição de RNA do vírus do herpes e da rubéola em células infectadas mostrou uma inibição respectivamente de 106 e 105 vezes, em relação ao controle. / In 2009, we demonstrated a potent antiviral in the hemolymph of L. obliqua. This purified protein has reduced the viral titer (TCID50 ml-1), measles and polio H1N1 157, 61 and 64 times. Recently, the antiviral protein expressed in baculovirus system, which reduced the title of the herpes virus in 106 times, rubella and EMC in 105 times. However, this expression system is very expensive and laborious. Thus, cloned and expressed this protein with antiviral activity in bacterial system. The antiviral sequence was cloned into pET28a expression vector. The resulting constructs were expressed in E. coli BL21 DE3 pLysS cells induced with 1.0 mM IPTG. Following expression, the bacterial pellet was sonicated and the material was purified by affinity and gel filtration. Was tested against EMC virus showing a protection of 104 times compared to the control. And using qPCR to determine the levels of viral RNA transcription of the herpes virus and rubella-infected cells showed an inhibition of respectively 106 and 105 times compared to the control.

The accessory protein Nef of human immunodeficiency virus type 1 (HIV-1) has long been known to enhance the infectivity of HIV-1 progeny virions. The multipass transmembrane proteins serine incorporator 3 (SERINC3) and SERINC5 were recently identified as novel antiviral proteins that restrict HIV-1 infectivity. Nef enhances HIV-1 infectivity by removing SERINCs from the plasma membrane, which prevents their incorporation into progeny HIV-1 virions. To exploit this potent intrinsic antiretroviral factor for potential therapy development, it is critical to explore the determinants in SERINC5 that govern its downregulation by Nef and its restriction on HIV-1 infectivity. Here I report that the ability to inhibit HIV-1 infectivity is conserved among vertebrate SERINC5 proteins, whereas the sensitivity to downregulation by Nef is not. However, a Nef-resistant SERINC5 became Nef-sensitive when its intracellular loop 4 (ICL4) was replaced by that of Nef-sensitive human SERINC5. Conversely, human SERINC5 became resistant to Nef when its ICL4 was replaced by that of a Nef-resistant SERINC5. In general, ICL4 regions from SERINCs that exhibited resistance to a given Nef conferred resistance to the same Nef when transferred to a sensitive SERINC, and vice versa. I demonstrate that human SERINC5 can be modified to restrict HIV-1 infectivity even in the presence of Nef. Moreover, by generating chimeras between SERINC5 and SERINC2, which does not exhibit antiretroviral activity, I demonstrate that SERINC5’s inhibitory function, unlike the sensitivity to Nef, requires the participation of more than one region. Helix 4 and extracellular loop 5 (ECL5) of SERINC5 are both required for the potent restriction of HIV-1 infectivity. In contrast, a large amino-terminal portion of SERINC5 is not required for its antiretroviral activity of SERINC5. The determinants in ECL5 disperse throughout the loop. Furthermore, the ECL5 of SERINC5 is a hotspot region that determines the Env-dependent antiretroviral activity of SERINC5.