Site-specific functionalization of proteins by organopalladium reactions

A new carbon-carbon bond has been regioselectively introduced into a target position (position 32 or 174) of the Ras protein by two types of organopalladium reactions (Mizoroki-Heck and Sonogashira reactions). Reaction conditions were screened by using a model peptide, and the stability of the Ras protein under the reaction conditions was examined by using the wild-type Ras protein. Finally, the iF-Ras proteins containing a 4-iodo-L-phenylalanine residue were subjected to organopalladium reactions with vinylated or propargylated biotin. Site-specific biotinylations of the Ras protein were confirmed by Western blot and LC-MS/MS.     

Soft materials with graphitic nanostructures

This review article focuses on our recent studies on novel soft materials consisting of carbon nanotubes. Single-walled carbon nanotubes, when suspended in imidazolium ion-based ionic liquids and ground in an agate mortar, form physical gels (bucky gels), where heavily entangled bundles of carbon nanotubes are exfoliated to give highly dispersed, much finer bundles. By using bucky gels, the first printable actuators that operate in air for a long time without any external electrolyte are developed. Furthermore, the use of polymerizable ionic liquids as the gelling media results in the formation of electroconductive polymer/nanotube composites with enhanced mechanical properties. The article also highlights a new family of nanotubular graphite, via self-assembly of amphiphilic hexabenzocoronene (HBC) derivatives. The nanotubes consist of a graphitic wall composed of a great number of pi-stacked HBC units and are electroconductive upon oxidation. The use of amphiphilic HBCs with functional groups results in the formation of nanotubes with various interesting properties.     

Possible Roles of Calreticulin in Uterine Decidualization and Receptivity in Rats and Humans

Previous in vitro studies have suggested that calreticulin (CALR), which is responsible for the folding and quality control of glycoproteins, may be associated with decidualization. However, its precise role in regulating decidualization has not been explored in vivo. Here, we used pregnant rat models to examine endometrial CALR expression during the peri-implantation period. We also examined whether polypectomy, a procedure that could ameliorate infertility, alters the endometrial expression levels of CALR and several implantation factors in women diagnosed as infertile. In rats, uterine CALR was expressed at a high level at the implantation site, and a marked increase in CALR expression was observed in decidual cells of normal pregnancy. In addition, endometrial CALR expression was enhanced by either administration of estradiol-17β in the delayed implantation rat model or the artificial induction of decidualization in the pseudopregnant rat. In cultured stromal cells, siRNA-mediated silencing of CALR inhibited the decidual stimulus-induced expression of prolactin, decidual/trophoblast prolactin-related protein, and connexin 43. In humans, the endometrial expression levels of the mRNAs encoding CALR and the implantation-related factor insulin-like growth factor binding protein (IGFBP)-7 tended to increase after polypectomy. The strongest positive correlation between expression levels before polypectomy was observed for IGFBP-7 and CALR, and the strength of this correlation increased after the surgery. Thus, endometrial CALR may play a role in the formation of decidua, and the polypectomy of infertile patients may result in the co-operative expression of endometrial factors, including CALR, that could enhance endometrial receptivity.     

Molecular modeling of single polypeptide chain of calcium-binding protein p26olf from dimeric S100B()

P26olf from olfactory tissue of frog, which may be involvedin olfactory transduction or adaptation, is a Ca²⁺-binding proteinwith 217 amino acids. The p26olf molecule contains two homologousparts consisting of the N-terminal half with amino acids 1–109and the C-terminal half with amino acids 110–217. Eachhalf resembles S100 protein with about 100 amino acids and containstwo helix–loop–helix Ca²⁺-binding structural motifsknown as EF-hands: a normal EF-hand at the C-terminus and apseudo EF-hand at the N-terminus. Multiple alignment of thetwo S100-like domains of p26olf with 18 S100 proteins indicatedthat the C-terminal putative EF-hand of each domain containsa four-residue insertion when compared with the typical EF-handmotifs in the S100 protein, while the N-terminal EF-hand ishomologous to its pseudo EF-hand. We constructed a three-dimensionalmodel of the p26olf molecule based on results of the multiplealignment and NMR structures of dimeric S100B(ßß)in the Ca²⁺-free state. The predicted structure of the p26olfsingle polypeptide chain satisfactorily adopts a folding patternremarkably similar to dimeric S100B(ßß). Each domainof p26olf consists of a unicornate-type four-helix bundle andthey interact with each other in an antiparallel manner formingan X-type four-helix bundle between the two domains. The twoS100-like domains of p26olf are linked by a loop with no sterichindrance, suggesting that this loop might play an importantrole in the function of p26olf. The circular dichroism spectraldata support the predicted structure of p26olf and indicatethat Ca²⁺-dependent conformational changes occur. Since theC-terminal putative EF-hand of each domain fully keeps the helix–loop–helixmotif having a longer Ca²⁺-binding loop, regardless of the four-residueinsertion, we propose that it is a new, novel EF-hand, althoughit is unclear whether this EF-hand binds Ca²⁺. P26olf is a newmember of the S100 protein family.     

Estimation and characterisation of major royal jelly proteins obtained from the honeybee Apis merifera

Royal jelly (RJ) contains many components, including proteins. We focused on major royal jelly proteins (MRJPs) under natural conditions, and attempted to determine the content ratios and molecular forms of MRJPs by size-exclusion HPLC, SDS–PAGE, 2-DE and MALDI TOF/TOF MS. Soluble RJ proteins were extracted by dialysis followed by several centrifugation techniques. Soluble RJ proteins were universally separated into five peaks (640 kDa, 280 kDa, 100 kDa, 72 kDa and 4.5 kDa) by size-exclusion HPLC on a Superose 12 column. Among these peaks, both the 280 kDa and 72 kDa peaks were major, but the intensity of the 280 kDa peak differed markedly among original RJ samples (n = 70). The main 280 kDa protein was separated into a 55 kDa band by reducing and non-reducing SDS–PAGE. This protein was also separated into multiple spots ranging from pH 4.2 to 6.5 by 2-DE. These spots were identified as MRJP 1 by MALDI TOF/TOF MS. From these results, MRJP 1 was thought to comprise an oligomer complex linked by non-covalent bonds under natural conditions. Another major protein, the 72 kDa peak on Superose 12 HPLC, was identified as MRJP 2.     

Fabrication of a polymer-coated silver hollow optical fiber with high performance

The techniques for fabricating a hollow optical fiber with an inner silver layer and a cyclic olefin polymer (COP) layer have been improved to reduce the surface roughness of these two layers. The loss spectrum was thereby drastically reduced over a wide wavelength range, from visible to near infrared. Optimization of the COP layer thickness resulted in low loss simultaneously at several key laser wavelengths. Infrared hollow fiber with low loss was developed for Er:YAG and Nd:YAG lasers. It can also deliver green and red pilot beams with low loss. Use of this fiber in therapeutic and pilot lasers should prove useful for research and development in laser medicine.     

Simultaneous enhanced photon capture and carrier generation in Si solar cells using Ge quantum dot photonic nanocrystals

We propose a novel solar cell structure with photonic nanocrystals coupled to quantum dots (QDs) for advanced management of photons and carriers. The photonic nanocrystals at the surface create an extra interaction between the photons and the QDs, which promotes light trapping. Photo-generated carriers can be efficiently transported by preparing vertically aligned QDs with electronic coupling. Implementation of the proposed structure was realized in crystalline Si solar cells with Ge QDs by development of a simple and practical formation method based on a wet chemical process without any lithography techniques. The wet process utilizes a periodically modulated etching rate induced by self-organized Ge QDs. The effectiveness of the proposed solar cell was demonstrated by the marked increase of the absolute conversion efficiency when compared with the control crystalline Si solar cells. It is found that light trapping by the photonic nanocrystals has a larger contribution to the efficiency improvement than the contributions from the carrier transport of the vertically aligned QDs.     

Amorphous nanosilicas induce consumptive coagulopathy after systemic exposure

We previously reported that well-dispersed amorphous nanosilicas with particle size 70 nm (nSP70) penetrate skin and produce systemic exposure after topical application. These findings underscore the need to examine biological effects after systemic exposure to nanosilicas. The present study was designed to examine the biological effects. BALB/c mice were intravenously injected with amorphous nanosilicas of sizes 70, 100, 300, 1000 nm and then assessed for survival, blood biochemistry, and coagulation. As a result, injection of nSP70 caused fatal toxicity, liver damage, and platelet depletion, suggesting that nSP70 caused consumptive coagulopathy. Additionally, nSP70 exerts procoagulant activity in vitro associated with an increase in specific surface area, which increases as diameter reduces. In contrast, nSP70-mediated procoagulant activity was absent in factor XII-deficient plasma. Collectively, we revealed that interaction between nSP70 and intrinsic coagulation factors such as factor XII, were deeply related to nSP70-induced harmful effects. In other words, it is suggested that if interaction between nSP70 and coagulation factors can be suppressed, nSP70-induced harmful effects may be avoided. These results would provide useful information for ensuring the safety of nanomaterials (NMs) and open new frontiers in biological fields by the use of NMs.     

Stacking faults in an epitaxially grown PbTiO3 thick film and their size distribution

The microstructure of an epitaxial PbTiO₃ thick film, grown on a SrRuO₃/SrTiO₃ substrate at 600 °C by pulsed-MOCVD method, was investigated by using transmission electron microscopy. A number of extrinsic or intrinsic stacking faults were observed in the epitaxial PbTiO₃ thick film and they were parallel to the (0 0 1) plane of the PbTiO₃. We also investigated the size distribution of these stacking faults. The width of these stacking faults along the [1 0 0] axis of the PbTiO₃ was very small, ranging from 2 to 13 nm. It was also revealed that the size distribution of stacking faults depends on the position in the film: near the surface, near the substrate, near threading dislocations, and near 90° domain boundaries.     

Flocculation of wall-deficient cells of Chlamydomonas reinhardtii mutant cw15 by calcium and methanol

Flocculation is a common and inexpensive method for harvesting algae from solution. After nitrogen starvation, it was shown that 83 ± 3% of the wall-deficient cells of the cw 15 mutant of Chlamydomonas reinhardtii flocculated from 12 mL samples within 15 min after the addition of 15 mM calcium chloride at pH 8.4. Only 24 ± 2% of the wildtype strain flocculated under these conditions, thus demonstrating how a simple mutation might facilitate process design. The data suggested that algae grown in waters with similar calcium concentrations (e.g. certain wastewaters) might be harvested through simple pH adjustment. It was also discovered that the addition of small amounts (<5% v/v) of methanol could significantly reduce the calcium needed to achieve flocculation. Within 15 min after addition of 12 mM calcium chloride and 4.6% (v/v) methanol, 83 ± 4% of cw15 cells flocculated. Methanol is fully recoverable by distillation, and its use might enable flocculation without further water salinization when media calcium concentrations fall short of 15 mM. It was further shown that substrates for and/or products of cellular growth affected flocculation adversely. Nearly 81% of cells flocculated from fresh medium compared to only 54% in spent medium.