Four Cysteine Residues Contribute to Homodimerization of Chicken Interleukin-2.

Interleukin-2 (IL-2) is a pleiotropic cytokine regulating the immune and nervous systems. Mammalian and bird IL-2s have different protein sequences, but perform similar functions. In the current study, two bands were detected by immunoblotting using an antibody against freshly purified chicken IL-2 (chIL-2). The molecular weight of the larger band was approximately twice as much of the chIL-2 monomer, although a chIL-2 complex or homodimer has never been reported. To explain this intriguing result, several dissociation reagents were used to examine the intermolecular forces between components of the proposed chIL-2 complex. It was found that intermolecular disulphide bond promotes homodimerization of chIL-2. Subsequently, mutation of Cys residues of chIL-2 revealed that mutation of all four Cys residues disrupted homodimerization, but a single, dual, or triple Cys mutation failed to disrupt homodimerization, suggesting that all four Cys residues on chIL-2 contribute to this dimerization. Functional analysis showed that both monomeric and dimeric chIL-2 consisting of either wild type or mutant chIL-2 were able to stimulate the expansion of CD4+ T cell in vivo or in vitro, and effectively bind to chIL-2 receptor. Overall, this study revealed that the recombinant chIL-2 purified from either Escherichia coli (E. coli) or Spodoptera frugiperda (Sf9) cells could homodimerize in vitro, with all four Cys residues on each chIL-2 protein contributing to this homodimerization, and dimerization and Cys mutation not impacting chIL-2 induced stimulation of chicken CD4+ T cells.

Design, synthesis and biological evaluation of pyridazino[3,4,5-de]quinazolin-3(2H)-one as a new class of PARP-1 inhibitors.

A series of pyridazino[3,4,5-de]quinazolin-3(2H)-one derivatives were designed and synthesized as PARP-1 inhibitors. Most of the synthesized compounds showed good inhibitory activities of PARP-1 and four of them achieved at the IC50 values ranging from 0.0914 µM to 0.244 µM. Two compounds, 1a and 1b, were further tested for their neuroprotective effect in the PC12 cell model injured by H2O2 and both of them exhibited excellent activities.