Two polymorphisms facilitate differences in plasticity between two chicken major histocompatibility complex class I proteins

Major histocompatibility complex class I molecules (MHC I) present peptides to cytotoxic T-cells at the surface of almost all nucleated cells. The function of MHC I molecules is to select high affinity peptides from a large intracellular pool and they are assisted in this process by co-factor molecules, notably tapasin. In contrast to mammals, MHC homozygous chickens express a single MHC I gene locus, termed BF2, which is hypothesised to have co-evolved with the highly polymorphic tapasin within stable haplotypes. The BF2 molecules of the B15 and B19 haplotypes have recently been shown to differ in their interactions with tapasin and in their peptide selection properties. This study investigated whether these observations might be explained by differences in the protein plasticity that is encoded into the MHC I structure by primary sequence polymorphisms. Furthermore, we aimed to demonstrate the utility of a complimentary modelling approach to the understanding of complex experimental data. Combining mechanistic molecular dynamics simulations and the primary sequence based technique of statistical coupling analysis, we show how two of the eight polymorphisms between BF2*15:01 and BF2*19:01 facilitate differences in plasticity. We show that BF2*15:01 is intrinsically more plastic than BF2*19:01, exploring more conformations in the absence of peptide. We identify a protein sector of contiguous residues connecting the membrane bound ?3 domain and the heavy chain peptide binding site. This sector contains two of the eight polymorphic residues. One is residue 22 in the peptide binding domain and the other 220 is in the ?3 domain, a putative tapasin binding site. These observations are in correspondence with the experimentally observed functional differences of these molecules and suggest a mechanism for how modulation of MHC I plasticity by tapasin catalyses peptide selection allosterically

Molecular determinants of chaperone interactions on MHC-I for folding and antigen repertoire selection.

The interplay between a highly polymorphic set of MHC-I alleles and molecular chaperones shapes the repertoire of peptide antigens displayed on the cell surface for T cell surveillance. Here, we demonstrate that the molecular chaperone TAP-binding protein related (TAPBPR) associates with a broad range of partially folded MHC-I species inside the cell. Bimolecular fluorescence complementation and deep mutational scanning reveal that TAPBPR recognition is polarized toward the α2 domain of the peptide-binding groove, and depends on the formation of a conserved MHC-I disulfide epitope in the α2 domain. Conversely, thermodynamic measurements of TAPBPR binding for a representative set of properly conformed, peptide-loaded molecules suggest a narrower MHC-I specificity range. Using solution NMR, we find that the extent of dynamics at "hotspot" surfaces confers TAPBPR recognition of a sparsely populated MHC-I state attained through a global conformational change. Consistently, restriction of MHC-I groove plasticity through the introduction of a disulfide bond between the α1/α2 helices abrogates TAPBPR binding, both in solution and on a cellular membrane, while intracellular binding is tolerant of many destabilizing MHC-I substitutions. Our data support parallel TAPBPR functions of 1) chaperoning unstable MHC-I molecules with broad allele-specificity at early stages of their folding process, and 2) editing the peptide cargo of properly conformed MHC-I molecules en route to the surface, which demonstrates a narrower specificity. Our results suggest that TAPBPR exploits localized structural adaptations, both near and distant to the peptide-binding groove, to selectively recognize discrete conformational states sampled by MHC-I alleles, toward editing the repertoire of displayed antigens

Dynamic Virtual Join Point Dispatch

Conceptually, join points are points in the execution of a program and advice is late-bound to them. We propose the notion of virtual join points that makes this concept explicit not only at a conceptual, but also at implementation level. In current implementations of aspect-oriented languages, binding is performed early, at deploy-time, and only a limited residual dispatch is executed. Current implementations fall in the categories of modifying the application code, modifying the meta-level of an application, or interacting with the application by means of events—the latter two already realizing virtual join points to some degree. We provide an implementation of an aspect-oriented execution environment that supports truly virtual join points and discuss how this approach also favors optimizations in the execution environment

Encapsulation of water insoluble drugs in mesoporous silica nanoparticles using supercritical carbon dioxide

Mesoporous silica nanoparticles MCM – 41 were synthesized with two dimensional hexagonal p6mm symmetry, high specific surface area(~ 980m2/g) narrow pore size and an average particle size of 186 nm. The produced nanoparticles were used to encapsulate carbamazepine through a supercritical carbon dioxide process combined with various organic solvents. Supercritical processing was found to provide increased drug encapsulation. The loaded MCM - 41 nanoparticles were analyzed using X–ray diffraction and differential scanning calorimetry (DSC) to investigate the crystalline state of the encapsulated carbamazepine and it was found to be dependent on the nature of the organic solvent. Carbamazepine showed increased dissolution rates under sink conditions. Viability studies of Caco – 2 cells demonstrated negligible cytotoxicity for the MCM–41 nanoparticles