(R)-2-(4-Phenylbutyl)dihydrobenzofuran derivatives as melatoninergic agents

(R)-2-(4-Phenylbutyl)dihydrobenzofuran derivatives (e.g., 3 and 4) were synthesized as novel melatoninergic ligands with significantly lower vasoconstrictive activity in vitro in the rat tail artery. Binding affinity assays were performed on cloned human MT1 and MT2 receptors stably expressed in NIH3T3 cells.     

Purification and properties of alkaline phosphatase of silkworm Bombyx mori

Alkaline phosphatase (AKP), from the succus entericus of silkworm, was purified using 10%–50% ammonium sulfate fractions, ion exchange chromatography of DEAE-Sepharose, and size exclusion chromatography of Sephacryl S-200. The purification fold was 464 times and specified activity was 3 936 U/mg. Optimum pH value of the phosphatase was 10.5, and was stable between pH 7.5 and 11. The optimum temperature of the phosphatase was 40°C and it was unstable over 50°C. K _m value of the phosphatase was 1.25 mmol/L. In a given condition, the phosphatase was selectively modified by PCMB, NBS, PMSF, TNBS, SUAN, DTT, BrAc, and IAc, the results indicate that PMSF, SUA, BrAc, IAc, and TNBS could obviously inhibit the activity of the phosphatase, and the degree of inhibition depended on the concentration of these reagents. There was little effect on the activity of phosphatase after treatment by PMSF, DTT, and NBT. We primarily conclude that mercapto and imidazole are essential for AKP from silkworm. Also, Lys residue and disulfide bands are necessary to protect the catalysis of the AKP. __________ Translated from Journal of Southwest Normal University (Natural Science), 2005, 30(5): 925–934 [译自: 西南师范大学学报 (自然科学版), 2005, 30(5): 925–934]     

Regulation of Skp2 Levels by the Pim-1 Protein Kinase

The Pim-1 protein kinase plays an important role in regulating both cell growth and survival and enhancing transformation by multiple oncogenes. The ability of Pim-1 to regulate cell growth is mediated, in part, by the capacity of this protein kinase to control the levels of the p27, a protein that is a critical regulator of cyclin-dependent kinases that mediate cell cycle progression. To understand how Pim-1 is capable of regulating p27 protein levels, we focused our attention on the SCF^Skp2 ubiquitin ligase complex that controls the rate of degradation of this protein. We found that expression of Pim-1 increases the level of Skp2 through direct binding and phosphorylation of multiple sites on this protein. Along with known Skp2 phosphorylation sites including Ser⁶⁴ and Ser⁷², we have identified Thr⁴¹⁷ as a unique Pim-1 phosphorylation target. Phosphorylation of Thr⁴¹⁷ controls the stability of Skp2 and its ability to degrade p27. Additionally, we found that Pim-1 regulates the anaphase-promoting complex or cyclosome (APC/C complex) that mediates the ubiquitination of Skp2. Pim-1 phosphorylates Cdh1 and impairs binding of this protein to another APC/C complex member, CDC27. These modifications inhibit Skp2 from degradation. Marked increases in Skp2 caused by these mechanisms lower cellular p27 levels. Consistent with these observations, we show that Pim-1 is able to cooperate with Skp2 to signal S phase entry. Our data reveal a novel Pim-1 kinase-dependent signaling pathway that plays a crucial role in cell cycle regulation.     

Optimization of γ-polyglutamic acid production byBacillus subtilis ZJU-7 using a surface-response methodology

The components of the media used to elicit the biosynthesis of poly-γ-glutamic acid (γ-PGA) byBacillus subtilis ZJU-7 were investigated, particularly the carbon and nitrogen sources. Of the 7 carbon sources investigated, sucrose induced the highest rate of γ-PGA productivity; among the nitrogen sources, tryptone had the best effect for γ-PGA production. A 2⁶⁻² fractional factorial design was used to screen factors that influence γ-PGA production significantly, and a central composite design was finally adopted to formulate the optimal medium. γ-PGA productivity improved approximately 2-fold when the optimal medium was used compared with the original nonoptimized medium, and volumetric productivity reached a maximum of 58.2 g/L after a 24-h cultivation period.     

Investigation on recombinant hirudin via oral route

The possibility for oral administration of peptide recombinant hirudin variant (rHV2-K47) as an anticoagulant agent was evaluated in several aspects. The proteolytic properties of rHV2-K47 and its stability during storage were examined by in vitro experiments. Radiolabeled rHV2-K47 was infused into the duodenum of rats and rHV2-K47 absorbed into serum was shown to be intact by electrophoresis pattern. The in vivo coagulation time of blood from mouse was prolonged significantly after oral administration of rHV2-K47. The bioavailability (F) of rHV2-K47 via oral route reached 10.11% in comparison with intravenous administration as gold standard. All the results suggested that rHV2-K47 could be delivered successfully via the oral route.     

A novel putative tyrosinase involved in periostracum formation from the pearl oyster (Pinctada fucata)

Tyrosinase (monophenol, L-DOPA: oxygen oxidoreductase, EC 1.14.18.1), a kind of copper-containing phenoloxidase, arouses great interests of scientists for its important role in periostracum formation. A cDNA clone encoding a putative tyrosinase, termed OT47 because of its estimated molecular mass of 47kDa, was isolated from the pearl oyster, Pinctada fucata. This novel tyrosinase shares similarity with the cephalopod tyrosinases and other type 3 copper proteins within two conserved copper-binding sites. RT-PCR analysis showed that OT47 mRNA was expressed only in the mantle edge. Further in situ hybridization analysis and tyrosinase activity staining revealed that OT47 was expressed at the outer epithelial cells of the middle fold, different from early histological results in Mercenaria mercenaria, suggesting a different model of periostracum secretion in P. fucata. Taken together, these results suggest that OT47 is most likely involved in periostracum formation. The identification and characterization of oyster tyrosinase also help to further understand the structural and functional properties of molluscan tyrosinase.     

A regulatory role for IL-10 receptor signaling in development and B cell help of T follicular helper cells in mice

IL -10 is widely accepted as a survival, proliferation, and differentiation factor for B cells. However, IL-10 deficiency accelerates disease progression as the result of autoantibody production in many autoimmune disease models. It was demonstrated that T follicular helper cells (T(FH) cells) play a key role in helping B cells that are secreting Abs. In this study, we demonstrated a regulatory role for IL-10R signaling on the development and B cell help function of T(FH) cells in vitro and in vivo. IL-1R subunit β-deficient (Il10rb(-/-)) Th cells were able to differentiate more readily into T(FH) cells, as well as secrete more IL-21 and IL-17 compared with wild-type Th cell-derived T(FH) cells. Increased IL-21 and IL-17 contributed to the enhanced B cell help functions of T(FH) cells. Further experiments demonstrated that IL-6 and IL-23 from dendritic cells in Il10rb(-/-) mice contributed to the differentiation of naive Th cells into T(FH) cells, as well as the generation of IL-21- and IL-17-producing T(FH) cells. Our results provide useful information for clarifying the immunoregulatory mechanisms associated with IL-10 deficiency in certain autoimmune disease models. This information could also be of benefit for the development of vaccines.