Germ Cell-free Protein Expression Research Articles

Evaluation of antibody responses to the early transcribed membrane protein family in Plasmodium vivax

BackgroundMalaria parasites form intracellular membranes that separate the parasite from the internal space of erythrocytes, and membrane proteins from the parasites are exported to the host via the membrane. In our previous study, Plasmodium vivax early transcribed membrane protein (PvETRAMP) 11.2, an intracellular membrane protein that is highly expressed in blood-stage parasites, was characterized as a highly immunogenic protein in P. vivax malaria patients. However, the other PvETRAMP family proteins have not yet been investigated. In this study, PvETRAMPs were expressed and evaluated to determine their immunological profiles.MethodsThe protein structure and amino acid alignment were carried out using bioinformatics analysis software. A total of six PvETRAMP family proteins were successfully expressed and purified using a wheat germ cell free protein expression system and the purified proteins were used for protein microarray and immunization of mice. The localization of the protein was determined with serum against PvETRAMP4. IgG subclasses were assessed from the immunized mice.ResultsIn silico analysis showed that P. vivax exhibits nine genes encoding the ETRAMP family. The ETRAMP family proteins are relatively small molecules with conserved structural features. A total of 6 recombinant ETRAMP proteins were successfully expressed and purified. The serum positivity of P. vivax malaria patients and healthy individuals was evaluated using a protein microarray method. Among the PvETRAMPs, ETRAMP4 showed the highest positivity rate of 62%, comparable to that of PvETRAMP11.2, which served as the positive control, and a typical export pattern of PvETRAMP4 was observed in the P. vivax parasite. The assessment of IgG subclasses in mice immunized with PvETRAMP4 showed high levels of IgG1 and IgG2b. PvETRAMP family proteins were identified and characterized as serological markers.ConclusionsThe relatively high antibody responses to PvETRAMP4 as well as the specific IgG subclasses observed in immunized mice suggest that the ETRAMP family is immunogenic in pathogens and can be used as a protein marker and for vaccine development.

Trends in wheat germ cell free protein expression system with an emphasis on up-scaling and industrial application

Cell-free translation systems have developed significantly over the last 2 decades and improvements in yield have resulted in their use for in vitro protein production in the laboratory. Such cell free systems enable the expression of cytotoxic, regulatory or unstable proteins that cannot be expressed in living cells. We compare the uses of different cell-free translation systems and review recent findings that support the possibility of scaling up and thereby enhancing the industrial application of wheat germ cell free protein expression. Keywords: Cell free biology, in vitro protein expression, scale up of translation, wheat germ extract

Trends in Wheat Germ Cell Free Protein Expression System with an Emphasis on Up-scaling and Industrial Application

Cell-free translation systems have developed significantly over the last 2 decades and improvements in yield have resulted in their use for in vitro protein production in the laboratory. Such cell free systems enable the expression of cytotoxic, regulatory or unstable proteins that cannot be expressed in living cells. We compare the uses of different cell-free translation systems and review recent findings that support the possibility of scaling up and thereby enhancing the industrial application of wheat germ cell free protein expression.