Transversal load distribution on bridge decks

An orthotropic rectangular plate is analysed. The plate has been considered simply supported in two opposite edges and general boundary conditions along the remainder edges. Matrix formulation, very convenient for programming on a digital computer, is used through the text. This technique is applied to an actual bridge deck and the results are compared with those obtained by means of the Guyon-Massonet-Rowe method

La transformación de la fachada en la arquitectura del siglo XX : evolución de los elementos arquitectónicos hacia el espacio

Esta tesis doctoral, bajo el título “La transformación de la fachada en la arquitectura del siglo XX. Evolución de los elementos arquitectónicos hacia el espacio único”, examina –siguiendo el método deductivo mediante investigación documental y de campo– los principales mecanismos proyectuales que han originado una transformación arquitectónica en la fachada. El término fachada, leído normalmente como sinónimo de arquitectura, ha sido analizado bajo taxonomías historiográficas, sociopolíticas, formales, compositivas o materiales. Más allá de estas clasificaciones, esta investigación determina cómo han evolucionado los elementos arquitectónicos hacia el espacio único, transformando con ello tanto la realidad física como el concepto de fachada. Proyectar, como diría Robert Venturi1. 1966, tanto desde el exterior al interior como desde el interior al exterior, produce las tensiones necesarias que ayudan a generar la arquitectura. De ahí la importancia del estudio de la transformación de la fachada como faccia, máscara, disfraz o sistema de representación, a la fachada como un diafragma activo, como un espacio de relación o como un límite donde se produce una experiencia arquitectónica con nuevos significados. El término fachada, del latín facies y del italiano facciata, fue creado como tal durante el siglo XVI, aunque como elemento arquitectónico es una realidad clásica y un concepto humanista reinventado en el Renacimiento. A principios del siglo XX la fachada se transforma radicalmente, con la aparición del vidrio plano de grandes dimensiones, y se despoja de su carga historicista. La transparencia literal hace desaparecer (negar) la fachada. La fachada se construye con nuevos materiales, nuevas tecnologías, y responde a un espacio global que espera a ser colonizado por una sociedad en constante mutación; un espacio que irá siendo cada vez más complejo a lo largo del siglo pasado y que tendrá como una de sus principales consecuencias un mayor grado de abstracción en la fachada. Para desarrollar con mayor precisión las intenciones de este trabajo se analiza la acepción de fachada en la que se encuadra la línea de investigación de la tesis doctoral. Siguiendo las indicaciones del Profesor Kenneth Frampton, se ha optado por dedicar precisamente a este análisis el primer capítulo, de precedentes, con el fin de hacer una relectura de la fachada barroca en clave contemporánea. La fachada barroca se estudia como un punto de inflexión, y origen de la transformación, frente al clasicismo de siglos precedentes. Se plantea por tanto como el primero de los mecanismos proyectuales motivo de estudio. Estos mecanismos han sido sometidos a una clasificación que responde a diversos tipos de espacio arquitectónico que han provocado la transformación de la fachada, analizando cada uno de ellos con un filtro bajo el cual se ha desarrollado un estudio pormenorizado. De esta manera se examinan la transformación compositiva de la fachada por medio del espacio dinámico (capítulo 00_precedentes) en San Carlo alle Quattro Fontane de Francesco Borromini, la transformación constructiva de la fachada por medio del espacio membrana (capítulo 01) en la Villa Tugendhat de Mies van der Rohe, la transformación diagramática de la fachada por medio del espacio pliegue (capítulo 02) en la terminal portuaria de Yokohama de FOA, la transformación tecnológica de la fachada por medio del espacio estructura (capítulo 03) en la mediateca de Sendai de Toyo Ito y, finalmente, la transformación fenomenológica de la fachada por medio del espacio múltiple (capítulo 04) en el pabellón de vidrio del Museo Toledo de SANAA. La investigación se completa con el análisis de una serie de textos que acompañan a cada uno de los capítulos, generando así un cuerpo de conocimiento global junto al análisis documental y las visitas de campo de cada uno de los casos de estudio considerados. ABSTRACT This doctoral thesis entitled “The Transformation of the Façade in Twentieth Century Architecture. The Evolution of Architectural Elements Towards a Single Space“ employs a deductive methodology incorporating both archival and field research, to examine the main design strategies that have given rise to the architectural transformation of the façade. The term façade, usually read as a synonym for architecture, has been analyzed under historiographic, sociopolitical, formal, compositive and material taxonomies. Far beyond these classifications, this study determines how architectural elements have evolved towards a single space, consequently transforming both the physical reality and the concept of façade. As Robert Venturi would say,1 to design from the inside-out and from the outside-in can create valid tensions that help generate architecture, hence the importance of this study of the transformation of the façade as a faccia, a mask, disguise or representation system, to the façade as an active diaphragm, a connective space or a limit within which architectural events with new meaning take place. The term fachada (Spanish for façade), from the Latin facies and the Italian facciata, appeared as such during the sixteenth century, although as an architectural element it is a classical ingredient and a humanist concept reinvented during the Renaissance. At the beginning of the 20th century, the façade is radically transformed with the introduction of large format plate glass, stripping it of its historicist content. Literal transparency causes the façade to disappear. The façade is built with new materials, new technologies, responding to a global space to be occupied by an everchanging society. It is a space that becomes increasingly more complex throughout the last century and which will have, as a consequence, a larger degree of abstraction in the façade. In order to more precisely focus this investigation, the meaning of the façade has been scrutinized. Following Professor Kenneth Frampton’s advice, the first chapter -Precedents- is precisely dedicated to this analysis, in order to understand the baroque façade read in a contemporary manner. The baroque façade has been studied as a turning point, the origin of the transformation of the façade, as compared to the classicism of previous centuries. For this reason it is considered the first design strategy to be studied. These strategeies have been sorted into a classification of architectural spaces that have caused the transformation of the façade, all of which having been analyzed using a methodology allowing for detailed study. Thus, this investigation examines the compositional transformation of the façade by means of a dynamic space (chapter 00: Precedents) in San Carlo alle Quattro Fontane by Francesco Borromini, the constructive transformation of the façade by means of a membrane space (chapter 01) in the Tugendhat Villa by Mies van der Rohe, the diagrammatic transformation of the façade by means of a folded space (chapter 02) in the Yokohama International Port Terminal by FOA, the technological transformation of the façade by means of a structure space (chapter 03) in the Sendai Mediathèque by Toyo Ito, and finally, the technological transformation of the façade by means of multiple space (chapter 04) in the Glass Pavilion of the Toledo Museum of Art by SANAA. The research is supplemented by the analysis of a series of texts presented alongside each chapter; creating a global body of knowledge together alongside the documentary analysis and on site analysis of each of the case studies considered.

Uncoupling of transfer of the presequence and unfolding of the mature domain in precursor translocation across the mitochondrial outer membrane

Translocation of mitochondrial precursor proteins across the mitochondrial outer membrane is facilitated by the translocase of the outer membrane (TOM) complex. By using site-specific photocrosslinking, we have mapped interactions between TOM proteins and a mitochondrial precursor protein arrested at two distinct stages, stage A (accumulated at 0°C) and stage B (accumulated at 30°C), in the translocation across the outer membrane at high resolution not achieved previously. Although the stage A and stage B intermediates were assigned previously to the forms bound to the cis site and the trans site of the TOM complex, respectively, the results of crosslinking indicate that the presequence of the intermediates at both stage A and stage B is already on the trans side of the outer membrane. The mature domain is unfolded and bound to Tom40 at stage B whereas it remains folded at stage A. After dissociation from the TOM complex, translocation of the stage B intermediate, but not of the stage A intermediate, across the inner membrane was promoted by the intermembrane-space domain of Tom22. We propose a new model for protein translocation across the outer membrane, where translocation of the presequence and unfolding of the mature domain are not necessarily coupled.

Amyloid fibril formation by an SH3 domain

The SH3 domain is a well characterized small protein module with a simple fold found in many proteins. At acid pH, the SH3 domain (PI3-SH3) of the p85α subunit of bovine phosphatidylinositol 3-kinase slowly forms a gel that consists of typical amyloid fibrils as assessed by electron microscopy, a Congo red binding assay, and x-ray fiber diffraction. The soluble form of PI3-SH3 at acid pH (the A state by a variety of techniques) from which fibrils are generated has been characterized. Circular dichroism in the far- and near-UV regions and 1H NMR indicate that the A state is substantially unfolded relative to the native protein at neutral pH. NMR diffusion measurements indicate, however, that the effective hydrodynamic radius of the A state is only 23% higher than that of the native protein and is 20% lower than that of the protein denatured in 3.5 M guanidinium chloride. In addition, the A state binds the hydrophobic dye 1-anilinonaphthalene-8-sulfonic acid, which suggests that SH3 in this state has a partially formed hydrophobic core. These results indicate that the A state is partially folded and support the hypothesis that partially folded states formed in solution are precursors of amyloid deposition. Moreover, that this domain aggregates into amyloid fibrils suggests that the potential for amyloid deposition may be a common property of proteins, and not only of a few proteins associated with disease.

Isolation of a myoglobin molten globule by selective cobalt(III)-induced unfolding

Reaction of the Schiff-base complex [Co(acetylacetonate-ethylenediimine)(NH3)2]+ with metmyoglobin at pH 6.5 yields a partially folded protein containing six Co(III) complexes. Although half of its α-helical secondary structure is retained, absorption and CD spectra indicate that the tertiary structure in both B-F and AGH domains is disrupted in the partially folded protein. In analogy to proton-induced unfolding, it is likely that the loss of tertiary structure is triggered by metal-ion binding to histidines. Cobalt(III)-induced unfolding of myoglobin is unique in its selectivity (other proteins are unaffected) and in allowing the isolation of the partially folded macromolecule (the protein does not refold or aggregate upon removal of free denaturant).

A model for the structure of the P domains in the subtilisin-like prohormone convertases

The proprotein convertases are a family of at least seven calcium-dependent endoproteases that process a wide variety of precursor proteins in the secretory pathway. All members of this family possess an N-terminal proregion, a subtilisin-like catalytic module, and an additional downstream well-conserved region of ≈150 amino acid residues, the P domain, which is not found in any other subtilase. The pro and catalytic domains cannot be expressed in the absence of the P domains; their thermodynamic instability may be attributable to the presence of large numbers of negatively charged Glu and Asp side chains in the substrate binding region for recognition of multibasic residue cleavage sites. Based on secondary structure predictions, we here propose that the P domains consist of 8-stranded β-barrels with well-organized inner hydrophobic cores, and therefore are independently folded components of the proprotein convertases. We hypothesize further that the P domains are integrated through strong hydrophobic interactions with the catalytic domains, conferring structural stability and regulating the properties and activity of the convertases. A molecular model of these interdomain interactions is proposed in this report.

Structural basis of an embryonically lethal single Ala → Thr mutation in the vnd/NK-2 homeodomain

The structural and DNA binding behavior is described for an analog of the vnd/NK-2 homeodomain, which contains a single amino acid residue alanine to threonine replacement in position 35 of the homeodomain. Multidimensional nuclear magnetic resonance, circular dichroism, and electrophoretic gel retardation assays were carried out on recombinant 80-aa residue proteins that encompass the wild-type and mutant homeodomains. The mutant A35T vnd/NK-2 homeodomain is unable to adopt a folded conformation free in solution at temperatures down to −5°C in contrast to the behavior of the corresponding wild-type vnd/NK-2 homeodomain, which is folded into a functional three-dimensional structure below 25°C. The A35T vnd/NK-2 binds specifically to the vnd/NK-2 target DNA sequence, but with an affinity that is 50-fold lower than that of the wild-type homeodomain. Although the three-dimensional structure of the mutant A35T vnd/NK-2 in the DNA bound state shows characteristic helix–turn–helix behavior similar to that of the wild-type homeodomain, a notable structural deviation in the mutant A35T analog is observed for the amide proton of leucine-40. The wild-type homeodomain forms an unusual i,i-5 hydrogen bond with the backbone amide oxygen of residue 35. In the A35T mutant this amide proton resonance is shifted upfield by 1.27 ppm relative to the resonance frequency for the wild-type analog, thereby indicating a significant alteration of this i,i-5 hydrogen bond.

Enhanced transcription factor access to arrays of histone H3/H4 tetramer⋅DNA complexes in vitro: Implications for replication and transcription

Defined model systems consisting of physiologically spaced arrays of H3/H4 tetramer⋅5S rDNA complexes have been assembled in vitro from pure components. Analytical hydrodynamic and electrophoretic studies have revealed that the structural features of H3/H4 tetramer arrays closely resemble those of naked DNA. The reptation in agarose gels of H3/H4 tetramer arrays is essentially indistinguishable from naked DNA, the gel-free mobility of H3/H4 tetramer arrays relative to naked DNA is reduced by only 6% compared with 20% for nucleosomal arrays, and H3/H4 tetramer arrays are incapable of folding under ionic conditions where nucleosomal arrays are extensively folded. We further show that the cognate binding sites for transcription factor TFIIIA are significantly more accessible when the rDNA is complexed with H3/H4 tetramers than with histone octamers. These results suggest that the processes of DNA replication and transcription have evolved to exploit the unique structural properties of H3/H4 tetramer arrays.

An optimal Mg2+ concentration for kinetic folding of the Tetrahymena ribozyme

Divalent metal ions, such as Mg2+, are generally required for tertiary structure formation in RNA. Although the role of Mg2+ binding in RNA-folding equilibria has been studied extensively, little is known about the role of Mg2+ in RNA-folding kinetics. In this paper, we explore the effect of Mg2+ on the rate-limiting step in the kinetic folding pathway of the Tetrahymena ribozyme. Analysis of these data reveals the presence of a Mg2+-stabilized kinetic trap that slows folding at higher Mg2+ concentrations. Thus, the Tetrahymena ribozyme folds with an optimal rate at 2 mM Mg2+, just above the concentration required for stable structure formation. These results suggest that thermodynamic and kinetic folding of RNA are cooptimized at a Mg2+ concentration that is sufficient to stabilize the folded form but low enough to avoid kinetic traps and misfolding.

Folding protein models with a simple hydrophobic energy function: The fundamental importance of monomer inside/outside segregation

The present study explores a “hydrophobic” energy function for folding simulations of the protein lattice model. The contribution of each monomer to conformational energy is the product of its “hydrophobicity” and the number of contacts it makes, i.e., E(h⃗, c⃗) = −Σi=1N cihi = −(h⃗.c⃗) is the negative scalar product between two vectors in N-dimensional cartesian space: h⃗ = (h1, … , hN), which represents monomer hydrophobicities and is sequence-dependent; and c⃗ = (c1, … , cN), which represents the number of contacts made by each monomer and is conformation-dependent. A simple theoretical analysis shows that restrictions are imposed concomitantly on both sequences and native structures if the stability criterion for protein-like behavior is to be satisfied. Given a conformation with vector c⃗, the best sequence is a vector h⃗ on the direction upon which the projection of c⃗ − c̄⃗ is maximal, where c̄⃗ is the diagonal vector with components equal to c̄, the average number of contacts per monomer in the unfolded state. Best native conformations are suggested to be not maximally compact, as assumed in many studies, but the ones with largest variance of contacts among its monomers, i.e., with monomers tending to occupy completely buried or completely exposed positions. This inside/outside segregation is reflected on an apolar/polar distribution on the corresponding sequence. Monte Carlo simulations in two dimensions corroborate this general scheme. Sequences targeted to conformations with large contact variances folded cooperatively with thermodynamics of a two-state transition. Sequences targeted to maximally compact conformations, which have lower contact variance, were either found to have degenerate ground state or to fold with much lower cooperativity.

Postgastrulation Smad2-deficient embryos show defects in embryo turning and anterior morphogenesis

SMAD2 is a member of the transforming growth factor β and activin-signaling pathway. To examine the role of Smad2 in postgastrulation development, we independently generated mice with a null mutation in this gene. Smad2-deficient embryos die around day 7.5 of gestation because of failure of gastrulation and failure to establish an anterior–posterior (A-P) axis. Expression of the homeobox gene Hex (the earliest known marker of the A-P polarity and the prospective head organizer) was found to be missing in Smad2-deficient embryos. Homozygous mutant embryos and embryonic stem cells formed mesoderm derivatives revealing that mesoderm induction is SMAD2 independent. In the presence of wild-type extraembryonic tissues, Smad2-deficient embryos developed beyond 7.5 and up to 10.5 days postcoitum, demonstrating a requirement for SMAD2 in extraembryonic tissues for the generation of an A-P axis and gastrulation. The rescued postgastrulation embryos showed malformation of head structures, abnormal embryo turning, and cyclopia. Our results show that Smad2 expression is required at several stages during embryogenesis.

Both apolipoprotein E and A-I genes are present in a nonmammalian vertebrate and are highly expressed during embryonic development

Apolipoprotein E (apoE) is associated with several classes of plasma lipoproteins and mediates uptake of lipoproteins through its ability to interact with specific cell surface receptors. Besides its role in cardiovascular diseases, accumulating evidence has suggested that apoE could play a role in neurodegenerative diseases, such as Alzheimer disease. In vertebrates, apoA-I is the major protein of high-density lipoprotein. ApoA-I may play an important role in regulating the cholesterol content of peripheral tissues through the reverse cholesterol transport pathway. We have isolated cDNA clones that code for apoE and apoA-I from a zebrafish embryo library. Analysis of the deduced amino acid sequences showed the presence of a region enriched in basic amino acids in zebrafish apoE similar to the lipoprotein receptor-binding region of human apoE. We demonstrated by whole-mount in situ hybridization that apoE and apoA-I genes are highly expressed in the yolk syncytial layer, an extraembryonic structure implicated in embryonic and larval nutrition. ApoE transcripts were also observed in the deep cell layer during blastula stage, in numerous ectodermal derivatives after gastrulation, and after 3 days of development in a limited number of cells both in brain and in the eyes. Our data indicate that apoE can be found in a nonmammalian vertebrate and that the duplication events, from which apoE and apoA-I genes arose, occurred before the divergence of the tetrapod and teleost ancestors. Zebrafish can be used as a simple and useful model for studying the role of apolipoproteins in embryonic and larval nutrition and of apoE in brain morphogenesis and regeneration.

Converting a transmembrane receptor to a soluble receptor: Recognition domain to effector domain signaling after excision of the transmembrane domain

The bacterial aspartate receptor was reconstructed to eliminate the transmembrane domain, thus connecting the recognition domain directly to the effector domain. The resulting soluble receptor folded correctly and was no longer an integral membrane protein. Upon aspartate binding, this soluble receptor was stabilized to a similar extent as that of the native receptor. Of interest, this soluble receptor retained the ability to signal from the recognition to the effector domain. This result defines more clearly the role of the membrane and transmembrane domains in signal transduction and suggests that some ligand-induced motions in receptor proteins do not require the membrane or transmembrane domain for information transmission.

Endoplasmic reticulum quality control of asialoglycoprotein receptor H2a involves a determinant for retention and not retrieval

The human asialoglycoprotein receptor H2a subunit contains a charged pentapeptide, EGHRG, in its ectodomain that is the only sequence absent from the H2b alternatively spliced variant. H2b exits the endoplasmic reticulum (ER) even when singly expressed, whereas H2a gives rise to a cleaved soluble secreted ectodomain fragment; uncleaved membrane-bound H2a molecules are completely retained and degraded in the ER. We have inserted the H2a pentapeptide into the sequence of the H1 subunit (H1i5), which caused complete ER retention but, unexpectedly, no degradation. This suggests that the pentapeptide is a determinant for ER retention not colocalizing in H2a with the determinant for degradation. The state of sugar chain processing and the ER localization of H1i5, which was unchanged at 15°C or after treatment with nocodazole, indicate ER retention and not retrieval from the cis-Golgi or the intermediate compartment. H1i5 folded similarly to H1, and both associated to calnexin. However, whereas H1 dissociated with a half time of 45 min, H1i5 remained bound to the chaperone for prolonged periods. The correct global folding of H2a and H1i5 and of other normal precursors and unassembled proteins and the true ER retention, and not exit and retrieval, suggest a difference in their quality control mechanism compared with that of misfolded proteins, which does involve retrieval. However, both pathways may involve calnexin.

Folding and aggregation of designed proteins

Protein aggregation is studied by following the simultaneous folding of two designed identical 20-letter amino acid chains within the framework of a lattice model and using Monte Carlo simulations. It is found that protein aggregation is determined by elementary structures (partially folded intermediates) controlled by local contacts among some of the most strongly interacting amino acids and formed at an early stage in the folding process.