Multivalent helix mimetics for PPI-inhibition.

The exploitation of multivalent ligands for the inhibition of protein-protein interactions has not yet been explored as a supramolecular design strategy. This is despite the fact that protein-protein interactions typically occur within the context of multi-protein complexes and frequently exploit avidity effects or co-operative binding interactions to achieve high affinity interactions. In this paper we describe preliminary studies on the use of a multivalent N-alkylated aromatic oligoamide helix mimetic for inhibition of p53/hDM2 and establish that protein dimerisation is promoted, rather than enhanced binding resulting from a higher effective concentration of the ligand. This journal i

Activation of NF-κB signalling by fusicoccin-induced dimerization

Chemically induced dimerization is an important tool in chemical biology for the analysis of protein function in cells. Here we report the use of the natural product fusicoccin (FC) to induce dimerization of 14-3-3-fused target proteins with proteins tagged to the C terminus (CT) of the H(+)-ATPase PMA2. To prevent nonproductive or detrimental interactions of the 14-3-3 proteins and CT fusions with endogenous cell proteins, their interaction surface was engineered to facilitate FC-induced dimerization exclusively between the introduced protein constructs. Live-cell imaging documented the reversible FC-induced translocation of 14-3-3 and CT to different cell compartments depending on localization sequences fused to their dimerization partner protein. The functionality of this system was demonstrated by the FC-induced importation of the NF-¿B-CT into the nucleus. In HeLa cells, FC-mediated dimerization of the NF-¿B-CT with a constitutively nuclear-localized 14-3-3 protein led to an NF-¿B-specific cellular response by inducing IL-8 secretion

Structure of a 14-3-3 sigma-YAP phosphopeptide complex at 1.15 a resolution

The 14-3-3 proteins are a class of eukaryotic acidic adapter proteins, with seven isoforms in humans. 14-3-3 proteins mediate their biological function by binding to target proteins and influencing their activity. They are involved in pivotal pathways in the cell such as signal transduction, gene expression, enzyme activation, cell division and apoptosis. The Yes-associated protein (YAP) is a WW-domain protein that exists in two transcript variants of 48 and 54 kDa in humans. By transducing signals from the cytoplasm to the nucleus, YAP is important for transcriptional regulation. In both variants, interaction with 14-3-3 proteins after phosphorylation of Ser127 is important for nucleocytoplasmic trafficking, via which the localization of YAP is controlled. In this study, 14-3-3 Sigma has been cloned, purified and crystallized in complex with a phosphopeptide from the YAP 14-3-3-binding domain, which led to a crystal that diffracted to 1.15 A resolution. The crystals belonged to space group C222(1), with unit-cell parameters a = 82.3, b = 112.1, c = 62.9 A

Structure of a 14-3-3 sigma-YAP phosphopeptide complex at 1.15 a resolution

The 14-3-3 proteins are a class of eukaryotic acidic adapter proteins, with seven isoforms in humans. 14-3-3 proteins mediate their biological function by binding to target proteins and influencing their activity. They are involved in pivotal pathways in the cell such as signal transduction, gene expression, enzyme activation, cell division and apoptosis. The Yes-associated protein (YAP) is a WW-domain protein that exists in two transcript variants of 48 and 54 kDa in humans. By transducing signals from the cytoplasm to the nucleus, YAP is important for transcriptional regulation. In both variants, interaction with 14-3-3 proteins after phosphorylation of Ser127 is important for nucleocytoplasmic trafficking, via which the localization of YAP is controlled. In this study, 14-3-3 Sigma has been cloned, purified and crystallized in complex with a phosphopeptide from the YAP 14-3-3-binding domain, which led to a crystal that diffracted to 1.15 A resolution. The crystals belonged to space group C222(1), with unit-cell parameters a = 82.3, b = 112.1, c = 62.9 A

Enrichment of chitosan hydrogels with perfluorodecalin promotes gelation and stem cell vitality

Thermosensitive injectable hydrogels for bone regeneration consisting of chitosan, sodium beta-glycerophosphate (Na-β-GP) and alkaline phosphatase (ALP) were enriched with oxygenated perfluorodecalin (PFD), a liquid hydrophobic perfluorochemical with high oxygen affinity, in order to improve cell growth on the hydrogels. Furthermore, influence of PFD concentration on hydrogel physicochemical properties relevant for bone regeneration, namely gelation speed, radiopacity and homogenicity, was investigated. Addtionally, ALP-mediated and non-ALP-mediated mineralization were evaluated by incubation in 0.1 M calcium glycerophosphate and simulated body fluid. 2% (w/v) chitosan hydrogels containing 2.5 mg/ml ALP were enriched with PFD at five concentrations, namely 0 (control), 0.069, 0.138, 0.207 and 0.276 ml/ml hydrogel, denoted A, B, C, D and E, respectively. Rheometrical investigations revealed that gelation speed increased with increasing PFD concentration. Micro-CT analysis revealed homogenicity of all sample groups except E and that radiopacity increased in the order B>C>A>D>E. ALP-mediated and non-ALP-mediated mineralization were not affected adversely by PFD. Growth of human adipose tissue-derived mesenchymal stem cells (ADSC) encapsulated in hydrogels was markedly higher in sample groups containing PFD, i.e. B–E. Hence, incorporation of oxygenated PFD can improve the suitability of hydrogels as bone regeneration materials