RESUMO
In humans, two endothelin receptors, ETa and ETb, are activated by three endogenous 21-mer cyclic peptides, ET-1, ET-2, and ET-3, which control various physiological processes, including vasoconstriction, vasodilation, and stimulation of cell proliferation. The first stage of this study it to produce a stable solubilized and purified receptor in a monodisperse state. This article is focused on the engineering, expression, purification, and characterization of the endothelin receptor B for subsequent structural and functional studies.
Assuntos
Receptor de Endotelina B/química , Receptor de Endotelina B/isolamento & purificação , Animais , Baculoviridae/genética , Compostos de Bifenilo/química , Western Blotting , Dipeptídeos/química , Antagonistas dos Receptores de Endotelina/química , Endotelinas/química , Engenharia Genética/métodos , Humanos , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Fragmentos de Peptídeos/química , Desnaturação Proteica , Estabilidade Proteica , Receptor de Endotelina B/genética , Receptor de Endotelina B/metabolismo , Proteínas Recombinantes de Fusão/química , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/isolamento & purificação , Proteínas Recombinantes de Fusão/metabolismo , Células Sf9 , TemperaturaRESUMO
This paper presents experimental data on the determination of the thickness of thylakoid membranes by small-angle neutron scattering. The thylakoids were isolated from spinach chloroplasts. The partial volume of proteins and lipids in the "washed" and "unwashed" membranes was estimated. It is shown that the thickness of thylakoid membranes, measured with this techniques depends on the way the membranes were separated. When isolated thylakoids by the standard method, the membrane thickness amounted to 75 A but if the extracted thylakoids were additionally washed with the isolation medium, the measured thickness was 50 A. In this case a significant decrease in the protein partial volume of the membrane was observed. The obtained results make it possible to explain numerous data on X-ray and small-angle neutron scattering by thylakoid membranes of different origins, proceeding from the assumption that all these membranes have a unified structure and consist of a stable core with a thickness of about 50 A, and layers of peripheral, weakly bound proteins with a thickness which may depends on the nature of the object under investigation and extracting conditions.