Co-evolution between pairs of residues in a multiple sequence alignment (MSA) of homologous proteins has long been proposed as an indicator of structural contacts. Recently, several methods, such as direct-coupling analysis (DCA) and MetaPSICOV, have been shown to achieve impressive rates of contact prediction by taking advantage of considerable sequence data. In this paper, we show that prediction success rates are highly sensitive to the structural definition of a contact, with more permissive definitions (i.e., those classifying more pairs as true contacts) naturally leading to higher positive predictive rates, but at the expense of the amount of structural information contributed by each contact. Thus, the remaining limitations of contact prediction algorithms are most noticeable in conjunction with geometrically restrictive contacts—precisely those that contribute more information in structure prediction. We suggest that to improve prediction rates for such “informative” contacts one could combine co-evolution scores with additional indicators of contact likelihood. Specifically, we find that when a pair of co-varying positions in an MSA is occupied by residue pairs with favorable statistical contact energies, that pair is more likely to represent a true contact. We show that combining a contact potential metric with DCA or MetaPSICOV performs considerably better than DCA or MetaPSICOV alone, respectively. This is true regardless of contact definition, but especially true for stricter and more informative contact definitions. In summary, this work outlines some remaining challenges to be addressed in contact prediction and proposes and validates a promising direction towards improvement.
Offshore concrete structures can be constructed in almost any country in the world using indigenous labour and materials. This can help oil companies meet agreed targets for the local content of a field development and in relation to technology transfer. However, construction in a developing region may be of concern particularly in terms of meeting the delivery schedule. The Malampaya concrete gravity substructure (CGS) project has demonstrated that offshore concrete structures can be constructed to the highest standards of safety and workmanship. The CGS was constructed in a remote part of the Philippines under a fixed price, lump sum contract faster than any previous similar project. During construction of the CGS there were no lost time incidents in 5 million man-hours worked. The paper describes those factors which contributed to this success. Data from the Malampaya project is used to provide guidance on how similar structures can be built in other developing regions around the world with confidence that the operator's objectives will be met. Background Attractions of Concrete Offshore Structures. Concrete gravity sub-structures (CGSs) have been in use for almost 30 years since the first platform, the Ecofisk Tank, was installed in the North Sea. The performance of the structures has been exemplary. Concrete has proved itself to be a highly durable, maintenance-free material suitable for the harshest of environments including ice-infested and highly seismic regions1. Dry-build CGSs are a more recent development 2. These structures are completed and pre-commissioned in an existing construction dock, a ship repair dry dock or a purpose-built casting basin, usually a simple "hole-in-the-ground". In general they have proved to be a more economical solution than jackets or floating systems in water depths up to about 150m especially when one or more of the following conditions applies:the field is remote and there are no existing pipeline export systems;in-field storage of oil or condensate is required;the platform is required to support heavy topsides;ground conditions are soft or unsuitable for piling. The structural form of CGSs is inherently suited to installation of the deck after emplacement of the CGS at the offshore location. A transportation barge supporting the deck is moored between the shafts and ballasted down until the load is permanently transferred to the support points on the top of the concrete shafts (Figure 1). This "float-over" installation technique has proved a cheaper alternative than either mating the deck and CGS in sheltered inshore water or using a heavy lift vessel at the offshore location. A more recent development is the concrete gravity tank (CGT) for offshore LNG facilities either for liquefaction or regassification (Figure 2). A number of schemes are now being actively considered for developments in Europe, West coast of Africa, Southeast Asia, Australia and North America. Concrete sub-sea storage tanks (CSSTs) are also under development as an economical, unmanned alternative to floating storage and offtake systems (Figure 3).
Cytokines are critical for guiding the differentiation of T lymphocytes to perform specialized tasks in the immune response. Developing strategies to manipulate cytokine-signaling pathways holds promise to program T cell differentiation toward the most therapeutically useful direction. Suppressor of cytokine signaling (SOCS) proteins are attractive targets, as they effectively inhibit undesirable cytokine signaling. However, these proteins target multiple signaling pathways, some of which we may need to remain uninhibited. SOCS3 inhibits IL-12 signaling but also inhibits the IL-2-signaling pathway. In this study, we use computational protein design based on SOCS3 and JAK crystal structures to engineer a mutant SOCS3 with altered specificity. We generated a mutant SOCS3 designed to ablate interactions with JAK1 but maintain interactions with JAK2. We show that this mutant does indeed ablate JAK1 inhibition, although, unexpectedly, it still coimmunoprecipitates with JAK1 and does so to a greater extent than with JAK2. When expressed in CD8 T cells, mutant SOCS3 preserved inhibition of JAK2-dependent STAT4 phosphorylation following IL-12 treatment. However, inhibition of STAT phosphorylation was ablated following stimulation with JAK1-dependent cytokines IL-2, IFN-a, and IL-21. Wild-type SOCS3 inhibited CD8 T cell expansion in vivo and induced a memory precursor phenotype. In vivo T cell expansion was restored by expression of the mutant SOCS3, and this also reverted the phenotype toward effector T cell differentiation. These data show that SOCS proteins can be engineered to fine-tune their specificity, and this can exert important changes to T cell biology.
Relating a protein's sequence to its conformation is a central challenge for both structure prediction and sequence design. Statistical contact potentials, as well as their more descriptive versions that account for side-chain orientation and other geometric descriptors, have served as simplistic but useful means of representing second-order contributions in sequence-structure relationships. Here we ask what happens when a pairwise potential is conditioned on the fully defined geometry of interacting backbones fragments. We show that the resulting structure-conditioned coupling energies more accurately reflect pair preferences as a function of structural contexts. These structure-conditioned energies more reliably encode native sequence information and more highly correlate with experimentally determined coupling energies. Clustering a database of interaction motifs by structure results in ensembles of similar energies and clustering them by energy results in ensembles of similar structures. By comparing many pairs of interaction motifs and showing that structural similarity and energetic similarity go hand-in-hand, we provide a tangible link between modular sequence and structure elements. This link is applicable to structural modeling, and we show that scoring CASP models with structuredconditioned energies results in substantially higher correlation with structural quality than scoring the same models with a contact potential. We conclude that structure-conditioned coupling energies are a good way to model the impact of interaction geometry on second-order sequence preferences.
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