Crystal structure of pathogenic Staphylococcus aureus lipase complex with the anti-obesity drug orlistat

抗肥満薬が黄色ブドウ球菌の病原因子を阻害するメカニズムを解明. 京都大学プレスリリース. 2020-03-31.Staphylococcus aureus lipase (SAL), a triacylglycerol esterase, is an important virulence factor and may be a therapeutic target for infectious diseases. Herein, we determined the 3D structure of native SAL, the mutated S116A inactive form, and the inhibitor complex using the anti-obesity drug orlistat to aid in drug development. The determined crystal structures showed a typical α/β hydrolase motif with a dimeric form. Fatty acids bound near the active site in native SAL and inactive S116A mutant structures. We found that orlistat potently inhibits SAL activity, and it covalently bound to the catalytic Ser116 residue. This is the first report detailing orlistat–lipase binding. It provides structure-based information on the production of potent anti-SAL drugs and lipase inhibitors. These results also indicated that orlistat can be repositioned to treat bacterial diseases

Development of a shutterless continuous rotation method using an X-ray CMOS detector for protein crystallography

A shutterless continuous rotation method using an X-ray complementary metal-oxide semiconductor (CMOS) detector has been developed for high-speed, precise data collection in protein crystallography. The new method and detector were applied to the structure determination of three proteins by multi- and single-wavelength anomalous diffraction phasing and have thereby been proved to be applicable in protein crystallography

Radically Different Thioredoxin Domain Arrangement of ERp46, an Efficient Disulfide Bond Introducer of the Mammalian PDI Family

SummaryThe mammalian endoplasmic reticulum (ER) contains a diverse oxidative protein folding network in which ERp46, a member of the protein disulfide isomerase (PDI) family, serves as an efficient disulfide bond introducer together with Peroxiredoxin-4 (Prx4). We revealed a radically different molecular architecture of ERp46, in which the N-terminal two thioredoxin (Trx) domains with positively charged patches near their peptide-binding site and the C-terminal Trx are linked by unusually long loops and arranged extendedly, forming an opened V-shape. Whereas PDI catalyzes native disulfide bond formation by the cooperative action of two mutually facing redox-active sites on folding intermediates bound to the central cleft, ERp46 Trx domains are separated, act independently, and engage in rapid but promiscuous disulfide bond formation during early oxidative protein folding. Thus, multiple PDI family members likely contribute to different stages of oxidative folding and work cooperatively to ensure the efficient production of multi-disulfide proteins in the ER

Architecture of the complete oxygen-sensing FixL-FixJ two-component signal transduction system

The symbiotic nitrogen-fixing bacterium Bradyrhizobium japonicum is critical to the agro-industrial production of soybean because it enables the production of high yields of soybeans with little use of nitrogenous fertilizers. The FixL and FixJ two-component system (TCS) of this bacterium ensures that nitrogen fixation is only stimulated under conditions of low oxygen. When it is not bound to oxygen, the histidine kinase FixL undergoes autophosphorylation and transfers phosphate from adenosine triphosphate (ATP) to the response regulator FixJ, which, in turn, stimulates the expression of genes required for nitrogen fixation. We purified full-length B. japonicum FixL and FixJ proteins and defined their structures individually and in complex using small-angle x-ray scattering, crystallographic, and in silico modeling techniques. Comparison of active and inactive forms of FixL suggests that intramolecular signal transduction is driven by local changes in the sensor domain and in the coiled-coil region connecting the sensor and histidine kinase domains. We also found that FixJ exhibits conformational plasticity not only in the monomeric state but also in tetrameric complexes with FixL during phosphotransfer. This structural characterization of a complete TCS contributes both a mechanistic and evolutionary understanding to TCS signal relay, specifically in the context of the control of nitrogen fixation in root nodules

Crystal structure of pathogenic Staphylococcus aureus lipase complex with the anti-obesity drug orlistat

抗肥満薬が黄色ブドウ球菌の病原因子を阻害するメカニズムを解明. 京都大学プレスリリース. 2020-03-31.Staphylococcus aureus lipase (SAL), a triacylglycerol esterase, is an important virulence factor and may be a therapeutic target for infectious diseases. Herein, we determined the 3D structure of native SAL, the mutated S116A inactive form, and the inhibitor complex using the anti-obesity drug orlistat to aid in drug development. The determined crystal structures showed a typical α/β hydrolase motif with a dimeric form. Fatty acids bound near the active site in native SAL and inactive S116A mutant structures. We found that orlistat potently inhibits SAL activity, and it covalently bound to the catalytic Ser116 residue. This is the first report detailing orlistat–lipase binding. It provides structure-based information on the production of potent anti-SAL drugs and lipase inhibitors. These results also indicated that orlistat can be repositioned to treat bacterial diseases

Size Limit on the Formation of Periodic Mesoporous Organosilicas (PMOs)

The decrease of the lattice size of periodic mesoporous organosilicas (PMOs) is one important goal in obtaining a microporous material for storage or adsorption of small molecules. To determine the influence of different synthesis parameters in the lattice size, here we performed in situ small-angle X-ray diffraction studies and show that a variation of the surfactant’s headgroup size is not directly followed by the lattice parameter of the resulting structure. We show that in the surfactant series of penta-, hexa-, hepta-, octa-, nona-, and decaethylene glycol monododecyl ether (C12(EO)n, n = 5, 6, 7, 8, 9, 10) the lattice size decreases between n = 5 and n = 8 and then increases, while the ordering of the materials is always cubic (space group Fd3m). This size effect is due to the ethylene oxide (EO) chain conformation that changes as the number of EO groups increases. Short ethylene oxide chains tend to have a so-called “zigzag” conformation while an increase of the chain length leads to a “Mäander” (coiling) conformation. Although this phenomenon is most commonly observed for chains consisting of more than 10 ethylene oxide units, we found a minimum PMO lattice size for 8 EO units and intermediate values for 6 and 7 EO units. The increase of the lattice parameter for more than 9 EO units is attributed to the increasing number of “Mäander” configurated EO units

Self-Sorting in the Formation of Metal–Organic Nanotubes: A Crucial Role of 2D Cooperative Interactions

A mixture of ferrocene-based tetratopic pyridyl ligands <b>FcL1</b> and <b>FcL2</b> undergoes self-sorting upon competitive coordination with AgBF₄, affording homomeric nanotubes <b>FcNT1</b> and <b>FcNT2</b> as a mixture. No mutual interference for the nanotubular growth occurred between <b>FcNT1</b> and <b>FcNT2</b> even when one of these ligands was used in large excess with respect to the other. 2D X-ray diffraction analysis of unidirectionally oriented nanotube samples, prepared by using the capillary technique, revealed that although <b>FcL1</b> as reported previously stacks helically in the resulting nanotube <b>FcNT1</b> <b>FcL2</b> prefers to stack with no discernible helical twist in <b>FcNT2</b>. Such a difference in their stacking geometries is most likely a major reason for why mixed-ligand metal–organic nanotubes are not constructed upon competitive coordination of <b>FcL1</b> and <b>FcL2</b> with AgBF₄