A folded and functional synthetic PA1b, an interlocked entomotoxic miniprotein

1-ACL (articles avec comité de lecture)PA1b (Pea Albumin 1, subunit b) is a hydrophobic, 37-amino acid niiniprotein isolated from pea seeds (Pivum sativum), crosslinked by three interlocked disulfide bridges, signature of the ICK (inhibitory cystine-knot) family. It acts as an entomotoxic factor against major insect pests in stored crops and vegetables, making it a promising bioinsecticide. Here we report an efficient and simple protocol for the production of large quantities of highly pure, biologically active synthetic PA1b. The features of PA1b oxidative refolding revealed the off-pathway products and competitive aggregation processes. The efficiency of the oxidative folding can be significantly improved by using hydrophobic alcoholic cosolvents and decreasing the temperature. The homogeneity of the synthetic oxidized PA1b was established by reversed-phase HPLC. The correct pairing of the three disulfide bridges, as well as the three-dimensional structure of synthetic PA1b was assessed by NMR. Synthetic PA1b binds to rnicrosomal proteins from Sitophilus oryzae with a Kd of 8 nM, a figure quite similar to that determined for PA1b extracted from its natural source. Moreover, the synthetic miniprotein was as potent as the extracted one towards the sensitive strains of weevils. Our findings will open the way to the production of PA1b analogues by chemical means to an in-depth understanding of the PA1b mechanism of action

In vitro wear testing of the PyroCarbon proximal interphalangeal joint replacement: Five million cycles of flexion and extension

Clinical results of the PyroCarbon proximal interphalangeal joint replacement are inconsistent with various complications reported. To address this, in vitro testing was conducted using finger joint simulators. Two PyroCarbon proximal interphalangeal prostheses were tested in a lubricant of dilute bovine serum to 5 × 106 cycles of flexion–extension (90°–30°) with dynamic forces of 10 N applied. At intervals of 3000 cycles testing ceased and a static load of 100 N was applied to simulate gripping. In addition, two ‘control’ prostheses were immersed alongside the test prostheses to account for lubricant absorption. Wear and roughness averages (Ra) were measured every 1 × 106 cycles. Minimal wear for all of the components was measured with a negligible increase in Ra for most of the components. One condyle of one component increased in Ra over the 5 × 106 cycles with a value above the recommended 50 nm. Unidirectional marks were visible on the condyle from micrographs, consistent with an abrasive wear mode

Vaccine-like Controlled-Release Delivery of an Immunomodulating Peptide to Treat Experimental Autoimmune Encephalomyelitis

The objective of this work is to use colloidal gel from alginate-chitosan-PLGA complex to deliver Ac-PLP-BPI-NH2-2 peptide in a controlled-release manner as a vaccine-like therapeutic to suppress experimental autoimmune encephalomyelitis (EAE) in the mouse model. Oppositely charged PLGA nanoparticles were prepared by a solvent diffusion method. The carboxyl group of the alginate and the amine group of the chitosan coated the nanoparticles with negative and positive charges, respectively. The peptide (Ac-PLP-BPI-NH2-2), designed to bind to MHC-II and ICAM-1 simultaneously, was formulated into the colloidal gel by physical mixture. Vaccine-like administration of the peptide-loaded colloidal gel (Ac-PLP-BPI-NH2-2-NP) was achieved by subcutaneous (s.c.) injection to EAE mice. Disease severity was measured using clinical scoring and percent change in body weight. Cytokine production was determined using the splenocytes from Ac-PLP-BPI-NH2-2-NP-treated mice and compared to that of controls. Ac-PLP-BPI-NH2-2-NP suppressed and delayed the onset of EAE as well as Ac-PLP-BPI-NH2-2 when delivered in a vaccine-like manner. IL-6 and IL-17 levels were significantly lower in the Ac-PLP-BPI-NH2-2-NP-treated mice compared to the mice group treated with blank colloidal gel, suggesting that the mechanism of suppression of EAE is due to a shift in the immune response away from Th17 production. The results of this study suggest that a one-time s.c. administration of Ac-PLP-BPI-NH2-2 formulated in a colloidal gel can produce long-term suppression of EAE by reducing Th17 proliferation