Techniques for the realization of ultrareliable spaceborne computers Interim scientific report

Reliability tests on spaceborne digital computer

Structure of a translocation signal domain mediating conjugative transfer by Type IV secretion systems

Relaxases are proteins responsible for the transfer of plasmid and chromosomal DNA from one bacterium to another during conjugation. They covalently react with a specific phosphodiester bond within DNA origin of transfer sequences, forming a nucleo-protein complex which is subsequently recruited for transport by a plasmid-encoded type IV secretion system. In previous work we identified the targeting translocation signals presented by the conjugative relaxase TraI of plasmid R1. Here we report the structure of TraI translocation signal TSA. In contrast to known translocation signals we show that TSA is an independent folding unit and thus forms a bona fide structural domain. This domain can be further divided into three sub-domains with striking structural homology with helicase sub-domains of the SF1B family. We also show that TSA is part of a larger vestigial helicase domain which has lost its helicase activity but not its single-stranded DNA binding capability. Finally, we further delineate the binding site responsible for translocation activity of TSA by targeting single residues for mutations. Overall, this study provides the first evidence that translocation signals can be part of larger structural scaffolds, overlapping with translocation-independent activities

Quantum Circuits for the Unitary Permutation Problem

We consider the Unitary Permutation problem which consists, given $n$ unitary gates $U_1, \ldots, U_n$ and a permutation $\sigma$ of $\{1,\ldots, n\}$, in applying the unitary gates in the order specified by $\sigma$, i.e. in performing $U_{\sigma(n)}\ldots U_{\sigma(1)}$. This problem has been introduced and investigated by Colnaghi et al. where two models of computations are considered. This first is the (standard) model of query complexity: the complexity measure is the number of calls to any of the unitary gates $U_i$ in a quantum circuit which solves the problem. The second model provides quantum switches and treats unitary transformations as inputs of second order. In that case the complexity measure is the number of quantum switches. In their paper, Colnaghi et al. have shown that the problem can be solved within $n^2$ calls in the query model and $\frac{n(n-1)}2$ quantum switches in the new model. We refine these results by proving that $n\log_2(n) +\Theta(n)$ quantum switches are necessary and sufficient to solve this problem, whereas $n^2-2n+4$ calls are sufficient to solve this problem in the standard quantum circuit model. We prove, with an additional assumption on the family of gates used in the circuits, that $n^2-o(n^{7/4+\epsilon})$ queries are required, for any $\epsilon >0$. The upper and lower bounds for the standard quantum circuit model are established by pointing out connections with the permutation as substring problem introduced by Karp.Comment: 8 pages, 5 figure

Structural and functional characterization of Pseudomonas aeruginosa CupB chaperones

Pseudomonas aeruginosa, an important human pathogen, is estimated to be responsible for,10% of nosocomial infections worldwide. The pathogenesis of P. aeruginosa starts from its colonization in the damaged tissue or medical devices (e. g. catheters, prothesis and implanted heart valve etc.) facilitated by several extracellular adhesive factors including fimbrial pili. Several clusters containing fimbrial genes have been previously identified on the P. aeruginosa chromosome and named cup [1]. The assembly of the CupB pili is thought to be coordinated by two chaperones, CupB2 and CupB4. However, due to the lack of structural and biochemical data, their chaperone activities remain speculative. In this study, we report the 2.5 A crystal structure of P. aeruginosa CupB2. Based on the structure, we further tested the binding specificity of CupB2 and CupB4 towards CupB1 (the presumed major pilus subunit) and CupB6 (the putative adhesin) using limited trypsin digestion and strep-tactin pull-down assay. The structural and biochemical data suggest that CupB2 and CupB4 might play different, but not redundant, roles in CupB secretion. CupB2 is likely to be the chaperone of CupB1, and CupB4 could be the chaperone of CupB4:CupB5:CupB6, in which the interaction of CupB4 and CupB6 might be mediated via CupB5

Unraveling the molecular basis of subunit specificity in P pilus assembly by mass spectrometry

P pili are multisubunit fibers essential for the attachment of uropathogenic Escherichia coli to the kidney. These fibers are formed by the noncovalent assembly of six different homologous subunit types in an array that is strictly defined in terms of both the number and order of each subunit type. Assembly occurs through a mechanism termed “donor-strand exchange (DSE)” in which an N-terminal extension (Nte) of one subunit donates a β-strand to an adjacent subunit, completing its Ig fold. Despite structural determination of the different subunits, the mechanism determining specificity of subunit ordering in pilus assembly remained unclear. Here, we have used noncovalent mass spectrometry to monitor DSE between all 30 possible pairs of P pilus subunits and their Ntes. We demonstrate a striking correlation between the natural order of subunits in pili and their ability to undergo DSE in vitro. The results reveal insights into the molecular mechanism by which subunit ordering during the assembly of this complex is achieved

Structure-function analysis reveals that the Pseudomonas aeruginosa Tps4 two-partner secretion system is involved in CupB5 translocation

Pseudomonas aeruginosa is a Gram-negative opportunistic bacterium, synonymous with cystic fibrosis patients, which can cause chronic infection of the lungs. This pathogen is a model organism to study biofilms: a bacterial population embedded in an extracellular matrix that provide protection from environmental pressures and lead to persistence. A number of Chaperone-Usher Pathways, namely CupA-CupE, play key roles in these processes by assembling adhesive pili on the bacterial surface. One of these, encoded by the cupB operon, is unique as it contains a nonchaperone-usher gene product, CupB5. Two-partner secretion (TPS) systems are comprised of a C-terminal integral membrane β-barrel pore with tandem N-terminal POTRA (POlypeptide TRansport Associated) domains located in the periplasm (TpsB) and a secreted substrate (TpsA). Using NMR we show that TpsB4 (LepB) interacts with CupB5 and its predicted cognate partner TpsA4 (LepA), an extracellular protease. Moreover, using cellular studies we confirm that TpsB4 can translocate CupB5 across the P. aeruginosa outer membrane, which contrasts a previous observation that suggested the CupB3 P-usher secretes CupB5. In support of our findings we also demonstrate that tps4/cupB operons are coregulated by the RocS1 sensor suggesting P. aeruginosa has developed synergy between these systems. Furthermore, we have determined the solution-structure of the TpsB4-POTRA1 domain and together with restraints from NMR chemical shift mapping and in vivo mutational analysis we have calculated models for the entire TpsB4 periplasmic region in complex with both TpsA4 and CupB5 secretion motifs. The data highlight specific residues for TpsA4/CupB5 recognition by TpsB4 in the periplasm and suggest distinct roles for each POTRA domain

Diethyl trans-2,5-bis­(4-methoxy­benzyl­sulfan­yl)-1,4-dimethyl-3,6-dioxopiperazine-2,5-carboxyl­ate

The title compound, C28H34N2O8S2, was synthesized as part of a project to develop synthetic routes to analogues of sporidesmins, a class of secondary metabolite produced by the filamentous fungi Chaetomium and Pithomyces sp. The complete molecule is generated by crystallographic inversion symmetry: the methoxy group is essentially coplanar with the benzene ring to which it is bonded, a mean plane fitted through the non-H atoms of the aromatic ring and the meth­oxy group having an r.m.s. deviation of 0.0140 Å. Similarly, the ester group is also essentially planar (r.m.s. deviation of a plane fitted through all non-H atoms is 0.0101 Å). There is only one independent C—H⋯O inter­action, which links together adjacent mol­ecules into a two-dimensional sheet in the bc plane

Desarrollo y evaluación mecánica de cubiertas poliméricas a base de nanocápsulas cargadas con aceite esencial de Thymus vulgaris.

A nivel mundial se estima que las pérdidas postcosecha de frutas y hortalizas frescas oscilan entre 15 y 85%. En los últimos años se han investigado las propiedades y aplicaciones del uso de recubrimientos y películas que mantengan la calidad de los alimentos y prolonguen la vida útil de estos. El aceite esencial de Thymus vulgaris han sido utilizado en cubiertas poliméricas por su actividad antibacteriana, sin embargo debido a su gran volatilidad, su uso se ha visto limitado por lo que es necesario su nanoencapsulación. Las NC obtenidas por la técnica de nanoprecipitación, fueron homogéneas y estables y se utilizaron para formar las cubiertas. Estas cubiertas fueron transparentes, homogéneas y resistentes al doblez. En base a sus propiedades mecánicas, se demostró que, las cubiertas formadas a base de NC, presentan un bajo porcentaje de elongación y rompimiento pero una adhesión elevada. Las cubiertas poliméricas obtenidas a base de NC cargadas con el aceite esencial de Thymus vulgaris, tienen un gran potencial como recubrimiento de frutas y hortalizas