RESUMO
Gene expression technology has made great progress with the continuous developments and researches of molecular biology. Though many systems to produce recombinant proteins have been studied, none of them is available so far to satisfy the needs completely. With the increasing demands of bioactive peptides and protein drugs, expression quantity and correct posttranslational folding and modifications are also needed under the circumstance which can make proteins more close to native conformation and higher activity and stability. Based on our previous work, we summarized the factors affecting the folding and modifications of recombinant proteins correctly from five aspects, including expression system and hosts, secretory expression, coexpression, fusion expression, and the culture conditions, as well as improvement strategies.
RESUMO
Heavy chain antibodies (HcAb) without light chains are naturally produced by camelids.The single domain antigen-binding fragment of HcAb is referred to as VHH or nanobody (Nb),which is the smallest antigen-binding entity at present.Several characteristics such as low molecular weight,high stability,and low immunogenicity make the use of nanobodies superior to the conventional antibodies.Currently,nanobodies are highly valuable antibodies for various applications,including fundamental research,diagnostics,and therapeutics.A variety of nanobodies are investigated in clinical researches.The recent progresses of heterologous expression of nanobodies in seven expression systems,including Gram-negative and positive bacteria,yeasts,filamentous fungi,insect cells,mammalian cells and plant cells were focused on,and the expression systems,hosts,characteristics of vectors,construction of vectors and yield of nanobodies were summarized.The strategies to increase the yield of nanobodies are discussed from molecular level,expression level and rational design.
RESUMO
Every day, new proteins are discovered and the need to understand its function arises. Human proteins have a special interest, because to know its role in the cell may lead to the design of a cure for a disease. In order to obtain such information, we need enough protein with a high degree of purity, and in the case of the human proteins, it is almost impossible to achieve this by working on human tissues. For that reason, the use of expression systems is needed. Bacteria, yeast, animals and plants have been genetically modified to produce proteins from different species. Even "cell-free" systems have been developed for that purpose. Here, we briefly review the options with their advantages and drawback, and the purification systems and analysis that can be done to gain understanding on the function and structure of the protein of interest.
Cada día, nuevas proteínas son descubiertas y surge la necesidad de caracterizarlas, siendo las de origen humano las que presentan un mayor interés. Conocer su función nos ayudará a entender padecimientos y diseñar una posible cura. Sin embargo, obtener suficiente cantidad de proteínas humanas en cantidad para llevar a cabo los análisis pertinentes, presenta una gran dificultad. Por tal razón, es necesario el uso de sistemas de expresión de proteínas heterólogas. Bacterias, levaduras, animales y plantas han sido modificados genéticamente para expresar proteínas de otras especies, e incluso sistemas in vitro han sido desarrollados para producir proteínas. En este artículo se revisan brevemente las opciones con sus ventajas y desventajas, así como las estrategias de purificación y los análisis que se pueden llevar a cabo para avanzar en el conocimiento de la función y estructura de la proteína de interés.