No abstract

The affinity of a ligand for a receptor on the cell surface will be influenced by the membrane composition. Herein, we evaluated the effects of differences in membrane fluidity, controlled by phospholipid composition, on the ligand binding activity of the G protein-coupled receptor human serotonin 2B. Using Nanodisc technology to control membrane properties, we performed biophysical analysis and employed molecular dynamics simulations to demonstrate that increased membrane fluidity shifted the equilibrium toward an active form of the receptor. Our quantitative study will enable development of more realistic in vitro drug discovery assays involving membrane-bound proteins such as G protein-coupled receptors.

Antibodies protect organisms from a huge variety of foreign antigens. Antibody diversity originates from both genetic and structural levels. Antigen recognition relies on complementarity between antigen-antibody interfaces. Recent methodological advances in structural biology and the accompanying rapid increase of the number of crystal structures of proteins have enabled atomic-level manipulation of protein structures to effect alterations in function. In this study, we explored the designability of electrostatic complementarity at an antigen-antibody interface on the basis of a crystal structure of the complex. We designed several variants with altered charged residues at the interface and characterized the designed variants by surface plasmon resonance, circular dichroism, differential scanning calorimetry, and molecular dynamics simulations. Both successes and failures of the structure-based design are discussed. The variants that compensate electrostatic interactions can restore the interface complementarity, enabling the cognate antigen-antibody binding. Retrospectively, we also show that these mutational effects could be predicted by the simulations. Our study demonstrates the importance of charged residues on the physical properties of this antigen-antibody interaction and suggests that computational approaches can facilitate design of antibodies that recognize a weakly immunogenic antigen.

The Wurtz type coupling of a mixture of meso-and dl-1,3-bis(chloromethylphenylsilyl)propanes with sodium metal afforded a mixture of cis-and trans-disilacyclopentanes (1a and 1b) in a ratio of 1:1. Pure 1a and 1b obtained by fractional distillation reacted with molecular oxygen in the presence of AIBN stereospecifically to give the respective cis-and trans-cyclic siloxanes (2a and 2b), with retention of configuration. The reactions of 1a and 1b with MCPBA produced cleanly 2a and 2b in high yields, respectively. The palladiumcatalyzed reactions of 1a and 1b with phenylacetylene gave stereospecifically cis-and trans-1,4-dimethyl-1,2,4-triphenyl-1,4-disilacyclohept-2-enes (3a and 3b), in high yields. Similar treatment of 1a and 1b with diphenylacetylene also proceeded with high stereospecificity to give cis-and trans-1,4-dimethyl-1,2,3,4-tetraphenyl-1,4-disilacyclohept-2-enes (4a and 4b), respectively. The results of an X-ray crystallographic study for products 4a and 4b are described.