Structural homologies with ATP- and folate-binding enzymes in the crystal structure of folylpolyglutamate synthetase

Folylpolyglutamate synthetase, which is responsible for the addition of a polyglutamate tail to folate and folate derivatives, is an ATP-dependent enzyme isolated from eukaryotic and bacterial sources, where it plays a key role in the retention of the intracellular folate pool. Here, we report the 2.4-Å resolution crystal structure of the MgATP complex of the enzyme from Lactobacillus casei. The structural analysis reveals that folylpolyglutamate synthetase is a modular protein consisting of two domains, one with a typical mononucleotide-binding fold and the other strikingly similar to the folate-binding enzyme dihydrofolate reductase. We have located the active site of the enzyme in a large interdomain cleft adjacent to an ATP-binding P-loop motif. Opposite this site, in the C domain, a cavity likely to be the folate binding site has been identified, and inspection of this cavity and the surrounding protein structure suggests that the glutamate tail of the substrate may project into the active site. A further feature of the structure is a well defined Ω loop, which contributes both to the active site and to interdomain interactions. The determination of the structure of this enzyme represents the first step toward the elucidation of the molecular mechanism of polyglutamylation of folates and antifolates.

Crystal structure of the ribosomal RNA domain essential for binding elongation factors

The structure of a 29-nucleotide RNA containing the sarcin/ricin loop (SRL) of rat 28 S rRNA has been determined at 2.1 Å resolution. Recognition of the SRL by elongation factors and by the ribotoxins, sarcin and ricin, requires a nearly universal dodecamer sequence that folds into a G-bulged cross-strand A stack and a GAGA tetraloop. The juxtaposition of these two motifs forms a distorted hairpin structure that allows direct recognition of bases in both grooves as well as recognition of nonhelical backbone geometry and two 5′-unstacked purines. Comparisons with other RNA crystal structures establish the cross-strand A stack and the GNRA tetraloop as defined and modular RNA structural elements. The conserved region at the top is connected to the base of the domain by a region presumed to be flexible because of the sparsity of stabilizing contacts. Although the conformation of the SRL RNA previously determined by NMR spectroscopy is similar to the structure determined by x-ray crystallography, significant differences are observed in the “flexible” region and to a lesser extent in the G-bulged cross-strand A stack.

Sla1p Is a Functionally Modular Component of the Yeast Cortical Actin Cytoskeleton Required for Correct Localization of Both Rho1p-GTPase and Sla2p, a Protein with Talin Homology

SLA1 was identified previously in budding yeast in a genetic screen for mutations that caused a requirement for the actin-binding protein Abp1p and was shown to be required for normal cortical actin patch structure and organization. Here, we show that Sla1p, like Abp1p, localizes to cortical actin patches. Furthermore, Sla1p is required for the correct localization of Sla2p, an actin-binding protein with homology to talin implicated in endocytosis, and the Rho1p-GTPase, which is associated with the cell wall biosynthesis enzyme β-1,3-glucan synthase. Mislocalization of Rho1p in sla1 null cells is consistent with our observation that these cells possess aberrantly thick cell walls.  Expression of mutant forms of Sla1p in which specific domains were deleted showed that the phenotypes associated with the full deletion are functionally separable. In particular, a region of Sla1p encompassing the third SH3 domain is important for growth at high temperatures, for the organization of cortical actin patches, and for nucleated actin assembly in a permeabilized yeast cell assay. The apparent redundancy between Sla1p and Abp1p resides in the C-terminal repeat region of Sla1p. A homologue of SLA1 was identified in Schizosaccharomyces pombe. Despite relatively low overall sequence homology, this gene was able to rescue the temperature sensitivity associated with a deletion of SLA1 in Saccharomyces cerevisiae.

X-ray structure of the human hyperplastic discs protein: An ortholog of the C-terminal domain of poly(A)-binding protein

The poly(A)-binding protein (PABP) recognizes the 3′ mRNA poly(A) tail and plays an essential role in eukaryotic translation initiation and mRNA stabilization/degradation. PABP is a modular protein, with four N-terminal RNA-binding domains and an extensive C terminus. The C-terminal region of PABP is essential for normal growth in yeast and has been implicated in mediating PABP homo-oligomerization and protein–protein interactions. A small, proteolytically stable, highly conserved domain has been identified within this C-terminal segment. Remarkably, this domain is also present in the hyperplastic discs protein (HYD) family of ubiquitin ligases. To better understand the function of this conserved region, an x-ray structure of the PABP-like segment of the human HYD protein has been determined at 1.04-Å resolution. The conserved domain adopts a novel fold resembling a right-handed supercoil of four α-helices. Sequence profile searches and comparative protein structure modeling identified a small ORF from the Arabidopsis thaliana genome that encodes a structurally similar but distantly related PABP/HYD domain. Phylogenetic analysis of the experimentally determined (HYD) and homology modeled (PABP) protein surfaces revealed a conserved feature that may be responsible for binding to a PABP interacting protein, Paip1, and other shared interaction partners.

Breaking down complex saproxylic communities: understanding sub-networks structure and implications to network robustness

Saproxylic insect communities inhabiting tree hollow microhabitats correspond with large food webs which simultaneously are constituted by multiple types of plant-animal and animal-animal interactions, according to the use of trophic resources (wood- and insect-dependent sub-networks), or to trophic habits or interaction types (xylophagous, saprophagous, xylomycetophagous, predators and commensals). We quantitatively assessed which properties of specialised networks were present in a complex networks involving different interacting types such as saproxylic community, and how they can be organised in trophic food webs. The architecture, interacting patterns and food web composition were evaluated along sub-networks, analysing their implications to network robustness from random and directed extinction simulations. A structure of large and cohesive modules with weakly connected nodes was observed throughout saproxylic sub-networks, composing the main food webs constituting this community. Insect-dependent sub-networks were more modular than wood-dependent sub-networks. Wood-dependent sub-networks presented higher species degree, connectance, links, linkage density, interaction strength, and were less specialised and more aggregated than insect-dependent sub-networks. These attributes defined high network robustness in wood-dependent sub-networks. Finally, our results emphasise the relevance of modularity, differences among interacting types and interrelations among them in modelling the structure of saproxylic communities and in determining their stability.

Highly organized structure in the non-coding region of the psbA minicircle from clade C Symbiodinium

The chloroplast genes of dinoflagellates are distributed among small, circular dsDNA molecules termed minicircles. In this paper, we describe the structure of the non-coding region of the psbA minicircle from Symbiodinium. DNA sequence was obtained from five Symbiodinium strains obtained from four different coral host species (Goniopora tenuidens, Heliofungia actiniformis, Leptastrea purpurea and Pocillopora damicornis), which had previously been determined to be closely related using LSU rDNA region D1/D2 sequence analysis. Eight distinct sequence blocks, consisting of four conserved cores interspersed with two metastable regions and flanked by two variable regions, occurred at similar positions in all strains. Inverted repeats (IRs) occurred in tandem or 'twin' formation within two of the four cores. The metastable regions also consisted of twin IRs and had modular behaviour, being either fully present or completely absent in the different strains. These twin IRs are similar in sequence to double-hairpin elements (DHEs) found in the mitochondrial genomes of some fungi, and may be mobile elements or may serve a functional role in recombination or replication. Within the central unit (consisting of the cores plus the metastable regions), all IRs contained perfect sequence inverses, implying they are highly evolved. IRs were also present outside the central unit but these were imperfect and possessed by individual strains only. A central adenine-rich sequence most closely resembled one in the centre of the non-coding part of Amphidinium operculatum minicircles, and is a potential origin of replication. Sequence polymorphism was extremely high in the variable regions, suggesting that these regions may be useful for distinguishing strains that cannot be differentiated using molecular markers currently available for Symbiodinium.

Modular alpha-helical mimetics with antiviral activity against respiratory syncitial virus

A 13-residue peptide sequence from a respiratory syncitial virus fusion protein was constrained in an alpha-helical conformation by fusing two back-to-back cyclic alpha-turn mimetics. The resulting peptide, Ac-(3 -> 7; 8 -> 12)-bicyclo-FP[KDEFD][KSIRD]V-NH2, was highly alpha-helical in water by CD and NMR spectroscopy, correctly positioning crucial binding residues (F488, I491, V493) on one face of the helix and side chain-side chain linkers on a noninteracting face of the helix. This compound displayed potent activity in both a recombinant fusion assay and an RSV antiviral assay (IC50 = 36 nM) and demonstrates for the first time that back-to-back modular alpha-helix mimetics can produce functional antagonists of important protein-protein interactions.

A modular holistic approach to prosody modelling for Standard Yorùbá speech synthesis

This paper presents a novel prosody model in the context of computer text-to-speech synthesis applications for tone languages. We have demonstrated its applicability using the Standard Yorùbá (SY) language. Our approach is motivated by the theory that abstract and realised forms of various prosody dimensions should be modelled within a modular and unified framework [Coleman, J.S., 1994. Polysyllabic words in the YorkTalk synthesis system. In: Keating, P.A. (Ed.), Phonological Structure and Forms: Papers in Laboratory Phonology III, Cambridge University Press, Cambridge, pp. 293–324]. We have implemented this framework using the Relational Tree (R-Tree) technique. R-Tree is a sophisticated data structure for representing a multi-dimensional waveform in the form of a tree. The underlying assumption of this research is that it is possible to develop a practical prosody model by using appropriate computational tools and techniques which combine acoustic data with an encoding of the phonological and phonetic knowledge provided by experts. To implement the intonation dimension, fuzzy logic based rules were developed using speech data from native speakers of Yorùbá. The Fuzzy Decision Tree (FDT) and the Classification and Regression Tree (CART) techniques were tested in modelling the duration dimension. For practical reasons, we have selected the FDT for implementing the duration dimension of our prosody model. To establish the effectiveness of our prosody model, we have also developed a Stem-ML prosody model for SY. We have performed both quantitative and qualitative evaluations on our implemented prosody models. The results suggest that, although the R-Tree model does not predict the numerical speech prosody data as accurately as the Stem-ML model, it produces synthetic speech prosody with better intelligibility and naturalness. The R-Tree model is particularly suitable for speech prosody modelling for languages with limited language resources and expertise, e.g. African languages. Furthermore, the R-Tree model is easy to implement, interpret and analyse.

Control of second-life hybrid battery energy storage system based on modular boost-multilevel buck converter

To fully utilize second-life batteries on the grid system, a hybrid battery scheme needs to be considered for several reasons: the uncertainty over using a single source supply chain for second-life batteries, the differences in evolving battery chemistry and battery configuration by different suppliers to strive for greater power levels, and the uncertainty of degradation within a second-life battery. Therefore, these hybrid battery systems could have widely different module voltage, capacity, and initial state of charge and state of health. In order to suitably integrate and control these widely different batteries, a suitable multimodular converter topology and an associated control structure are required. This paper addresses these issues proposing a modular boost-multilevel buck converter based topology to integrate these hybrid second-life batteries to a grid-tie inverter. Thereafter, a suitable module-based distributed control architecture is introduced to independently utilize each converter module according to its characteristics. The proposed converter and control architecture are found to be flexible enough to integrate widely different batteries to an inverter dc link. Modeling, analysis, and experimental validation are performed on a single-phase modular hybrid battery energy storage system prototype to understand the operation of the control strategy with different hybrid battery configurations.

An analytical investigation into shear wall and slab interconnections between reinforced concrete modules for high -rise buildings utilising modular construction under extreme seismic and wind loading

This research investigates a new structural system utilising modular construction. Five-sided boxes are cast on-site and stacked together to form a building. An analytical model was created of a typical building in each of two different analysis programs utilising the finite element method (Robot Millennium and ETABS). The pros and cons of both Robot Millennium and ETABS are listed at several key stages in the development of an analytical model utilising this structural system. Robot Millennium was initially utilised but created an analytical model too large to be successfully run. The computation requirements were too large for conventional computers. Therefore Robot Millennium was abandoned in favour of ETABS, whose more simplistic algorithms and assumptions permitted running this large computation model. Tips are provided as well as pitfalls signalled throughout the process of modelling such complex buildings of this type. ^ The building under high seismic loading required a new horizontal shear mechanism. This dissertation has proposed to create a secondary floor that ties to the modular box through the use of gunwales, and roughened surfaces with epoxy coatings. In addition, vertical connections necessitated a new type of shear wall. These shear walls consisted of waffled external walls tied through both reinforcement and a secondary concrete pour. ^ This structural system has generated a new building which was found to be very rigid compared to a conventional structure. The proposed modular building exhibited a period of 1.27 seconds, which is about one-fifth of a conventional building. The maximum lateral drift occurs under seismic loading with a magnitude of 6.14 inches which is one-quarter of a conventional building's drift. The deflected shape and pattern of the interstorey drifts are consistent with those of a coupled shear wall building. In conclusion, the computer analysis indicate that this new structure exceeds current code requirements for both hurricane winds and high seismic loads, and concomitantly provides a shortened construction time with reduced funding. ^

New dimeric supramolecular structure of mixed (phthalocyaninato)(porphyrinato)-europium(III) sandwiches : preparation and spectroscopic characteristics

The mixed double-decker Eu\[Pc(15C5)4](TPP) (1) was obtained by base-catalysed tetramerisation of 4,5-dicyanobenzo-15-crown-5 using the half-sandwich complex Eu(TPP)(acac) (acac = acetylacetonate), generated in situ, as the template. For comparative studies, the mixed triple-decker complexes Eu2\[Pc(15C5)4](TPP)2 (2) and Eu2\[Pc(15C5)4]2(TPP) (3) were also synthesised by the raise-by-one-story method. These mixed ring sandwich complexes were characterised by various spectroscopic methods. Up to four one-electron oxidations and two one-electron reductions were revealed by cyclic voltammetry (CV) and differential pulse voltammetry (DPV). As shown by electronic absorption and infrared spectroscopy, supramolecular dimers (SM1 and SM3) were formed from the corresponding double-decker 1 and triple-decker 3 in the presence of potassium ions in MeOH/CHCl3.