Sparse and incomplete factorial matrices to screen membrane protein 2D crystallization.

Electron crystallography is well suited for studying the structure of membrane proteins in their native lipid bilayer environment. This technique relies on electron cryomicroscopy of two-dimensional (2D) crystals, grown generally by reconstitution of purified membrane proteins into proteoliposomes under conditions favoring the formation of well-ordered lattices. Growing these crystals presents one of the major hurdles in the application of this technique. To identify conditions favoring crystallization a wide range of factors that can lead to a vast matrix of possible reagent combinations must be screened. However, in 2D crystallization these factors have traditionally been surveyed in a relatively limited fashion. To address this problem we carried out a detailed analysis of published 2D crystallization conditions for 12 ß-barrel and 138 α-helical membrane proteins. From this analysis we identified the most successful conditions and applied them in the design of new sparse and incomplete factorial matrices to screen membrane protein 2D crystallization. Using these matrices we have run 19 crystallization screens for 16 different membrane proteins totaling over 1300 individual crystallization conditions. Six membrane proteins have yielded diffracting 2D crystals suitable for structure determination, indicating that these new matrices show promise to accelerate the success rate of membrane protein 2D crystallization.

A New Method to Determine the Transmembrane Conformation of Substrates in Intramembrane Proteolysis by Deep-UV Resonance Raman Spectroscopy.

We present a new method based on deep-UV resonance Raman spectroscopy to determine the backbone conformation of intramembrane protease substrates. The classical amide vibrational modes reporting on the conformation of just the transmembrane region of the substrate can be resolved from solvent exchangeable regions outside the detergent micelle by partial deuteration of the solvent. In the presence of isotopically triple-labeled intramembrane protease, these amide modes can be accurately measured to monitor the transmembrane conformation of the substrate during intramembrane proteolysis.

Prenatal development of adenohypophyseal cell types, ovary, and uterus of dromedary camel.

The somatotrophic and thyrotrophic cells were originally developed in the adenohypophysis from undifferentiated chromophobic cells at the foetal age of 4-8 weeks. The follicle-stimulating hormone cells appeared at a foetal age of 24-28 weeks. Prolactin and luteinizing hormone cells were first seen at of foetal age levels of 32-36 and 40 weeks. The germinal epithelium of flattened and cuboidal cells in the ovary was changed to cubical and columnar types, 24-28 weeks of foetal age. The sex cords appeared at 8-12 weeks of foetal age. The primary and secondary follicles were observed at 8-12 and 20-24 weeks of foetal age, respectively. Follicular cavity was noticed at 32-36 weeks, while large follicles encapsulated by follicular sheath were seen in the 40th week of foetal age. Some ovarian follicles degenerated during or after their development. Surface epithelium in the uterus changed from cubical to columnar type at 8-12 weeks of foetal age. The endometrium was clearly distinguished from the myometrium at 16-20 weeks of foetal age. The uterine glands appeared at 16-20 weeks as short and straight tubular glands, and they began to branch in the 40th week of foetal age.