RESUMEN
Photosystem II subunit S (PsbS) is a membrane protein that plays an exclusive role in non-photochemical quenching for photoprotection of plants under high-light conditions. The activation mechanism of PsbS and its pH-induced conformational changes are currently unknown. For structural investigation of PsbS, effective synthesis of PsbS with selective isotope or electron-spin labels or non-natural amino acids incorporated would be a great asset. Here we present cell-free (CF) expression as a successful method for in vitro production of PsbS that would allow such incorporation. The addition of several detergents, liposomes and lipid nanodiscs was tested for achieving soluble CF expression of PsbS. We have optimized the CF method to yield soluble PsbS of â¼500â¯ng/µl using a continuous-exchange method at 30⯰C, along with a successful purification and refolding of PsbS in n-Dodecyl ß-D-maltoside (ß-DM) detergent. We expect that the presented protocols are transferrable for in vitro expression of other membrane proteins of the Light-Harvesting Complex family.
Asunto(s)
Complejo de Proteína del Fotosistema II/genética , Técnicas Biosensibles , Glucósidos/química , Concentración de Iones de Hidrógeno , Lípidos/química , Liposomas/química , Nanoestructuras/química , Procesos Fotoquímicos , Complejo de Proteína del Fotosistema II/química , Conformación Proteica , Pliegue de Proteína , SolubilidadRESUMEN
Normal values for the flow at a transnasal pressure of 150 Pa were established with active anterior rhinomanometry (with decongestion) in a group of 33 normal subjects. These values were used to detect abnormalities in a group of 193 patients whose septum anatomy had been evaluated with rhinoscopy. About 25% of the rhinoscopically normal patients were found to have significantly low ("abnormal") flow values on one side. The same was true for patients with a small septal deviation restricted to one anatomical area. An abnormal flow was measured in about 35% of the patients with a moderate (restricted) septal deviation. In the patients whose septal deviation was not restricted to one anatomical area, about 45% had an abnormal flow. The highest detection rate was about 80% in patients with major deviations in the region of the vestibule and the valve. Such deviations were found only in a minority of the patients with complaints of nasal obstruction, which limits the importance of rhinomanometric evaluation in clinical practice.