The properties of vesicle membranes prepared from 16:0-SM, 16:0-DHSM, or DPPC were characterized using steady-state and time-resolved fluorescence spectroscopy and different fluorescent reporter molecules. The acyl-chain region was probed using free and phospholipid-bound 1,6-diphenyl-1,3,5-hexatriene. 16:0-DHSM was found to be the more ordered than both DPPC and 16:0-SM 5 degrees C below and above melting temperature. Interfacial properties of the phospholipid bilayers were examined using 6-dodecanoyl-2-dimethyl-aminonaphthalene (Laurdan), 6-propionyl-2-dimethyl-amino-naphthalene (Prodan), and dansyl-PE. Laurdan and Prodan reported that the two sphingomyelin (SM) membrane interfaces were clearly different from the DPPC membrane interface, whereas the two SM membrane interfaces had more similar properties (both in gel and liquid-crystalline phase). Prodan partition studies showed that membrane resistance to Prodan partitioning increased in the order: 16:0-SM < DPPC < 16:0-DHSM. The degree to which dansyl-PE is exposed to water reflects the structural properties of the membrane-water interface. By comparing the lifetime of dansyl-PE in water and deuterium oxide solution, we could show that the degree to which the dansyl moiety was exposed to water in the membranes increased in the order: 16:0-SM < DPPC < 16:0-DHSM. In conclusion, this study has shown that DHSM forms more ordered bilayers than acyl-chain matched SM or phosphatidylcholine, even in the liquid-crystalline state.
Human vascular adhesion protein-1 (VAP-1) is a membrane-bound multifunctional glycoprotein with both adhesive and enzymatic properties. The protein belongs to the copper-containing amine oxidase (CAO) family, which use 2,4,5-trihydroxyphenylalanine quinone as a cofactor. Here, the crystallization and preliminary X-ray analysis of a mammalian CAO, human VAP-1, is reported. The protein was expressed in Chinese hamster ovary cells as a full-length form with an N-terminal transmembrane region and multiple glycosylation sites. Hexagonal crystals with unit-cell parameters a = b = 225.9, c = 218.7 A, alpha = beta = 90, gamma = 120 degrees were obtained using the vapour-diffusion method. Data from three different crystals were collected at 100 K using synchrotron radiation and were processed to 3.2 A resolution with 95.9% completeness and an R(merge) of 19.6%.
1275The atoms deviate by a maximum of 0.022 A from the plane of the whole molecule; thus it is planar within experimental error to a significance level of about 0.5 according to a X 2 test. (The isotropic e.s.d.'s for the atomic positions are 0.012-0.016,/%.) The benzene rings A and C are planar to within +0.003 and _+0.012 A, respectively, and the furan rings B and D are planar to within +0.009 and -+0.007A, respectively.The molecules are stacked along the short a axis, as indicated by the packing diagram in Fig. 2. All intermolecular contacts correspond to normal van der Waals distances. The distance of closest approach between adjacent molecules is 3.74 (1)A. In spite of the non-interacting distances, the packing is very dense, as reflected in the high D x of 1.664 (1)Mg m -3. The packing coefficient, i.e. the ratio between the volume occupied by the four molecules and the cell volume, is 0.75.The authors are indebted to Jan-Eric Berg for stimulating discussions and also to Professor Peder Kierkegaard for his continued interest in this work. The crystals used were kindly supplied by Professor Holger Erdtman and Dr Hans-Erik H6gberg.The present research program has received financial support from the Swedish Natural Science Research Council.References BANERJEE, A. (1973). Acta Cryst. B29, 2070B29, -2074B29, . BANERJEE, A. (1974 1.391 (1) Mg m -3, /z(Cu Kct) = 0.73 mm -1, F(000) = 536. The final R value is 0.040 for 1082 data collected at ambient temperature using Cu Ka radiation (2 = 1.5418 A) and with /net > 4tr(Inet). The molecule is significantly non-planar with an angle of 4.1 (3) ° between the outer benzene rings.
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