Allosteric Activation of the Par-6 PDZ Via a Partial Unfolding Transition

Proteins exist in a delicate balance between the native and unfolded states, where thermodynamic stability may be sacrificed to attain the flexibility required for efficient catalysis, binding, or allosteric control. Partition-defective 6 (Par-6) regulates the Par polarity complex by transmitting a GTPase signal through the Cdc42/Rac interaction binding PSD-95/Dlg/ZO-1 (CRIB-PDZ) module that alters PDZ ligand binding. Allosteric activation of the PDZ is achieved by local rearrangement of the L164 and K165 side chains to stabilize the interdomain CRIB:PDZ interface and reposition a conserved element of the ligand binding pocket. However, microsecond to millisecond dynamics measurements revealed that L164/K165 exchange requires a larger rearrangement than expected. The margin of thermodynamic stability for the PDZ domain is modest (∼3 kcal/mol) and further reduced by transient interactions with the disordered CRIB domain. Measurements of local structural stability revealed that tertiary contacts within the PDZ are disrupted by a partial unfolding transition that enables interconversion of the L/K switch. The unexpected participation of partial PDZ unfolding in the allosteric mechanism of Par-6 suggests that native-state unfolding may be essential for the function of other marginally stable proteins

Some (dis)assembly required: partial unfolding in the Par-6 allosteric switch

Allostery is commonly described as a functional connection between two distant sites in a protein, where a binding event at one site alters affinity at the other. Here, we review the conformational dynamics that encode an allosteric switch in the PDZ domain of Par-6, which is a scaffold protein that organizes other proteins into a complex required to initiate and maintain cell polarity. NMR measurements revealed that the PDZ domain samples an evolutionarily conserved unfolding intermediate allowing rearrangement of two adjacent loop residues that control ligand binding affinity. Cdc42 binding to Par-6 creates a novel interface between the PDZ domain and the adjoining CRIB motif that stabilizes the high-affinity PDZ conformation. Thermodynamic and kinetic studies suggest that partial PDZ unfolding is an integral part of the Par-6 switching mechanism. The Par-6 CRIB-PDZ module illustrates two important structural aspects of protein evolution: the interface between adjacent domains in the same protein can give rise to allosteric regulation, and thermodynamic stability may be sacrificed to increase the sampling frequency of an unfolding intermediate required for conformational switching.National Institutes of Health (U.S.) (grants R01 AI058072, R56 AI013225, and S10 RR024665

Regret, Portfolio Choice, and Guarantees in Defined Contribution Schemes

We model how asset allocation decisions in a defined contribution (DC) pension plan might vary with participants’ attitudes about risk and regret. We show that anticipated disutility from regret can have a potent effect on investment choices. Compared to a risk-averse investor, the investor who takes regret into account will hold more stock when the equity premium is low but less stock when the equity premium is high. We also assess how regret can influence a DC plan participant’s view of rate-of-return guarantees, as measured by his willingness-to-pay. We find that regret increases the regret-averse investor’s willingness to pay for a guarantee when the portfolio is relatively risky but decreases it when the portfolio is relatively safe

Nmr Solution Structure Of Plastocyanin From The Photosynthetic Prokaryote, Prochlorothrix Hollandica

The solution structure of a divergent plastocyanin (PC) from the photosynthetic prokaryote Prochlorothrix hollandica was determined by homonuclear H-1 NMR spectroscopy. Nineteen structures were calculated from 1222 distance restraints, yielding a family of structures having an average rmsd of 0.42 +/- 0.08 Angstrom, for backbone atoms and 0.71 +/- 0.07 Angstrom for heavy atoms to the mean structure. No distance constraint was violated by more than 0.26 Angstrom in the structure family. Despite the low number of conserved residues shared with other PC homologues, the overall folding pattern of P. hollandica PC is similar to other PCs, in that the protein forms a two-sheet beta-barrel tertiary structure. The greatest variability among the backbone structures is seen in the loop region from residues 47-60. The differences seen in the P. hollandica PC homologue likely arise due to a small deletion of 2-4 residues compared to the PC consensus; this yields a less extended loop containing a short alpha-helix from residues Ala52-Leu55. Additionally, the protein has an altered hydrophobic patch thought to be important in binding reaction partners. Whereas the backbone structure is very similar within the loops of the hydrophobic region, the presence of two unique residues (Tyr12 and Pro14) yields a structurally different hydrophobic surface likely important in binding P. hollandica Photosystem I

Evolution of an ancient protein function involved in organized multicellularity in animals.

To form and maintain organized tissues, multicellular organisms orient their mitotic spindles relative to neighboring cells. A molecular complex scaffolded by the GK protein-interaction domain (GKPID) mediates spindle orientation in diverse animal taxa by linking microtubule motor proteins to a marker protein on the cell cortex localized by external cues. Here we illuminate how this complex evolved and commandeered control of spindle orientation from a more ancient mechanism. The complex was assembled through a series of molecular exploitation events, one of which - the evolution of GKPID's capacity to bind the cortical marker protein - can be recapitulated by reintroducing a single historical substitution into the reconstructed ancestral GKPID. This change revealed and repurposed an ancient molecular surface that previously had a radically different function. We show how the physical simplicity of this binding interface enabled the evolution of a new protein function now essential to the biological complexity of many animals

Altered drug susceptibility during host adaptation of a <i>Plasmodium falciparum</i> strain in a non-human primate model

Infections with Plasmodium falciparum, the most pathogenic of the Plasmodium species affecting man, have been reduced in part due to artemisinin-based combination therapies. However, artemisinin resistant parasites have recently emerged in South-East Asia. Novel intervention strategies are therefore urgently needed to maintain the current momentum for control and elimination of this disease. In the present study we characterize the phenotypic and genetic properties of the multi drug resistant (MDR) P. falciparum Thai C2A parasite strain in the non-human Aotus primate model, and across multiple passages. Aotus infections with C2A failed to clear upon oral artesunate and mefloquine treatment alone or in combination, and ex vivo drug assays demonstrated reduction in drug susceptibility profiles in later Aotus passages. Further analysis revealed mutations in the pfcrt and pfdhfr loci and increased parasite multiplication rate (PMR) across passages, despite elevated pfmdr1 copy number. Altogether our experiments suggest alterations in parasite population structure and increased fitness during Aotus adaptation. We also present data of early treatment failures with an oral artemisinin combination therapy in a pre-artemisinin resistant P. falciparum Thai isolate in this animal model

Rapid sulfurisation of highly branched isoprenoid (HBI) alkenes in sulfidic Holocene sediments from Ellis Fjord, Antarctica

Author Posting. © Elsevier B.V., 2007. This is the author's version of the work. It is posted here by permission of Elsevier B.V. for personal use, not for redistribution. The definitive version was published in Organic Geochemistry 38 (2007): 128-139, doi:10.1016/j.orggeochem.2006.08.003.Samples of particulate organic matter from the water column and anoxic Holocene sediment layers from the Small Meromictic Basin (SMB) in Ellis Fjord (eastern Antarctica) were analyzed to study the early incorporation of reduced inorganic sulfur species into highly branched isoprenoid (HBI) alkenes. HBIs were not detected in the water column samples from austral winter, whereas compounds containing the C25 HBI skeleton were abundant in all analyzed Holocene sediment layers. The structure of the C25:2 HBI alkene together with its enriched stable carbon isotopic composition suggest that the HBI alkene is produced by a diatom or diatoms probably belonging to the Navicula genus present in the sea-ice which covers the area most of the year. Within just 500 years of deposition, all of the HBI alkene was sulfurised. A mixture of products was formed, including components tentatively identified as a C25 HBI thiane and three S-containing dimers composed of two C25:1 HBI skeletons linked together by a sulfide bond. Most of the HBI alkene, however, was converted to polar S-containing compounds. The observed reaction rate for sulfurisation the C25:2 HBI alkene is the highest observed so far in natural systems. Sterols and other lipids known to be prone to sulfurisation were only minimally sulfurised under these depositional conditions. The reason for this is presently unclear.Funding for the collection of the sediment and water samples (by MJLC and CW) was provided by ASAC grant 1166 to JKV. This work was further supported by a grant from the Netherlands Organization for Scientific Research (NWO; Netherlands Antarctic Research Proposals 851.20.006 to JSSD)

Crave-Out: A Distraction/Motivation Mobile Game to Assist in Smoking Cessation

BACKGROUND: Smoking is still the number one preventable cause of death. Cravings-an intense desire or longing for a cigarette-are a major contributor to quit attempt failure. New tools to help smokers\u27 manage their cravings are needed. OBJECTIVE: To present a case study of the development process and testing of a distraction/motivation game (Crave-Out) to help manage cravings. METHODS: We used a phased approach: in Phase 1 (alpha testing), we tested and refined the game concept, using a Web-based prototype. In Phase 2 (beta testing), we evaluated the distraction/motivation potential of the mobile game prototype, using a prepost design. After varying duration of abstinence, smokers completed the Questionnaire of Smoking Urge-Brief (QSU-Brief) measurement before and after playing Crave-Out. Paired t tests were used to compare pregame and postgame QSU-Brief levels. To test dissemination potential, we released the game on the Apple iTunes App Store and tracked downloads between December 22, 2011, and May 5, 2014. RESULTS: Our concept refinement resulted in a multilevel, pattern memory challenge game, with each level increasing in difficulty. Smokers could play the game as long as they wanted. At the end of each level, smokers were provided clear goals for the next level and rewards (positive reinforcement using motivational tokens that represented a benefit of quitting smoking). Negative reinforcement was removed in alpha testing as smokers felt it reminded them of smoking. Measurement of QSU-Brief (N=30) resulted in a pregame mean of 3.24 (SD 1.65) and postgame mean of 2.99 (SD 1.40) with an overall decrease of 0.25 in cravings (not statistically significant). In a subset analysis, the QSU-Brief decrease was significant for smokers abstinent for more than 48 hours (N=5) with a pregame mean of 2.84 (SD 1.16) and a postgame mean of 2.0 (SD 0.94; change=0.84; P =.03). Between December 22, 2011, and May 29, 2014, the game was downloaded 3372 times from the App-Store, with 1526 smokers visiting the online resource www.decide2quit.org linked to the game. CONCLUSIONS: Overall, playing the game resulted in small, but nonsignificant decreases in cravings, with changes greater for those had already quit for more than 48 hours. Lessons learned can inform further development. Future research could incorporate mHealth games in multicomponent cessation interventions. TRIAL REGISTRATION: Clinicaltrials.gov NCT00797628; https://clinicaltrials.gov/ct2/show/NCT00797628 (Archived by WebCite at http://www.webcitation.org/6hbJr6LWG)

A long‑term precision agriculture system sustains grain profitability

After two decades of availability of grain yield-mapping technology, long-term trends in field-scale profitability for precision agriculture (PA) systems and conservation practices can now be assessed. Field-scale profitability of a conventional or ‘business-as-usual’ system with an annual corn (Zea mays L.)-soybean (Glycine max [L.]) rotation and annual tillage was assessed for 11 years on a 36 ha field in central Missouri during 1993 to 2003. Following this, a ‘precision agriculture system’ (PAS) with conservation practices was implemented for the next 11 years to address production, profit and environmental concerns. The PAS was multifaceted and temporally dynamic. It included no-till, cover crops, crop rotation changes, site-specific N and variable-rate or zonal P, K and lime. Following a recent evaluation of differences in yield and yield variability, this research compared profitability of the two systems. Results indicated that PAS sustained profits in the majority (97%) of the field without subsidies for cover crops or payments for enhanced environmental protection. Profit was only lower with PAS in a drainage channel where no-till sometimes hindered soybean stands and wet soils caused wheat (Triticum aestivum L.) disease. Although profit gains were not realized after 11 years of PA and conservation practices, this system sustained profits. These results should help growers gain confidence that PA and conservation practices will be successful

Molecular assays for antimalarial drug resistance surveillance: A target product profile.

Antimalarial drug resistance is a major constraint for malaria control and elimination efforts. Artemisinin-based combination therapy is now the mainstay for malaria treatment. However, delayed parasite clearance following treatment with artemisinin derivatives has now spread in the Greater Mekong Sub region and may emerge or spread to other malaria endemic regions. This spread is of great concern for malaria control programmes, as no alternatives to artemisinin-based combination therapies are expected to be available in the near future. There is a need to strengthen surveillance systems for early detection and response to the antimalarial drug resistance threat. Current surveillance is mainly done through therapeutic efficacy studies; however these studies are complex and both time- and resource-intensive. For multiple common antimalarials, parasite drug resistance has been correlated with specific genetic mutations, and the molecular markers associated with antimalarial drug resistance offer a simple and powerful tool to monitor the emergence and spread of resistant parasites. Different techniques to analyse molecular markers associated with antimalarial drug resistance are available, each with advantages and disadvantages. However, procedures are not adequately harmonized to facilitate comparisons between sites. Here we describe the target product profiles for tests to analyse molecular markers associated with antimalarial drug resistance, discuss how use of current techniques can be standardised, and identify the requirements for an ideal product that would allow malaria endemic countries to provide useful spatial and temporal information on the spread of resistance