Solution Structure of the Squash Aspartic Acid Proteinase Inhibitor (SQAPI) and Mutational Analysis of Pepsin Inhibition

The squash aspartic acid proteinase inhibitor (SQAPI), a proteinaceous proteinase inhibitor from squash, is an effective inhibitor of a range of aspartic proteinases. Proteinaceous aspartic proteinase inhibitors are rare in nature. The only other example in plants probably evolved from a precursor serine proteinase inhibitor. Earlier work based on sequence homology modeling suggested SQAPI evolved from an ancestral cystatin. In this work, we determined the solution structure of SQAPI using NMR and show that SQAPI shares the same fold as a plant cystatin. The structure is characterized by a four-strand anti-parallel β-sheet gripping an α-helix in an analogous manner to fingers of a hand gripping a tennis racquet. Truncation and site-specific mutagenesis revealed that the unstructured N terminus and the loop connecting β-strands 1 and 2 are important for pepsin inhibition, but the loop connecting strands 3 and 4 is not. Using ambiguous restraints based on the mutagenesis results, SQAPI was then docked computationally to pepsin. The resulting model places the N-terminal strand of SQAPI in the S′ side of the substrate binding cleft, whereas the first SQAPI loop binds on the S side of the cleft. The backbone of SQAPI does not interact with the pepsin catalytic Asp³²–Asp²¹⁵ diad, thus avoiding cleavage. The data show that SQAPI does share homologous structural elements with cystatin and appears to retain a similar protease inhibitory mechanism despite its different target. This strongly supports our hypothesis that SQAPI evolved from an ancestral cystatin.     

γ-Butyrobetaine hydroxylase catalyses a Stevens type rearrangement

γ-Butyrobetaine hydroxylase (BBOX) is a 2-oxoglutarate and Fe(II)-dependent oxygenase that catalyses the final step of L-carnitine biosynthesis in animals. BBOX catalyses the oxidation of 3-(2,2,2-trimethylhydrazinium)propionate (THP), a clinically used BBOX inhibitor, to form multiple products including 3-amino-4-(methyamino)butanoic acid (AMBA), which is proposed to be formed via a Stevens type rearrangement mechanism. We report the synthesis of AMBA and confirm that it is a product of the BBOX catalysed oxidation of THP. AMBA reacts with formaldehyde, which is produced enzymatically by BBOX, to give a cyclic adduct.     

Nitrogen content, amino acid composition and digestibility of fungi from a nutritional perspective in animal mycophagy

Fungi comprise a major part of the diet of many animals. Even so, the nutritional value of fungi has been much debated, with some arguing that fungi are nutritionally poor. However, the chemical composition of fungi and of the biology of the animals that eat them are not well understood, particularly in reference to amino acid (AA) composition of fungi and digestibility of fungal protein. We analysed fibre, total nitrogen (N), available N, and AA contents and measured in vitro digestibility of a wide range of epigeous and hypogeous fungi collected in Australia and the USA to test three hypotheses: (i) fungi are nutritionally poor because they contain few nutrients or are otherwise of low digestibility, (ii) fungi vary substantially in their nutritional composition; and (iii) animals can counter this variable quality by eating diverse taxa. Resultant data indicate many fungi are a reasonable source of AAs and digestible nitrogen. However, they vary highly between species in AA content, and the protein has a poor balance of digestible AAs. This helps explain why many mycophagous animals eat a wide array of fungi and often have digestive strategies to cope with fungi, such as foregut fermentation. Another common strategy is to supplement the diet with high quality protein, such as insect protein. Accordingly, evaluating nutritional value of fungi requires consideration of physiology of the animal species and their whole diet.     

Desert Truffles of the Australian Outback: Ecology,Ethnomycology, and Taxonomy

Desert Truffles of the Australian Outback: Ecology, Ethnomycology, and Taxonomy. The Aborigines of central Australia have traditionally used desert truffles as food. Truffle hunting in the desert requires substantial ecological knowledge, as truffles occur sporadically and only with adequate and properly distributed rainfall as well as the presence of necessary soil conditions and mycorrhizal hosts. Truffles are hunted primarily by women, who look for cracks or humps in the soil caused by expansion of the truffles, which are then extracted with digging sticks. The truffles are typically eaten raw or baked or roasted in ashes. Seven truffle species are recorded from the Australian Outback, including three that have been only recently described.     

N-(3-fluoro-4-(2-arylthieno[3,2-b]pyridin-7-yloxy)phenyl)-2-oxo-3-phenylimidazolidine-1-carboxamides: A novel series of dual c-Met/VEGFR2 receptor tyrosine kinase inhibitors

A series of N-(3-fluoro-4-(2-arylthieno[3,2-b]pyridin-7-yloxy)phenyl)-2-oxo-3-phenylimidazolidine-1-carboxamides targeting c-Met and VEGFR2 tyrosine kinases was designed and synthesized. The compounds were potent against these two enzymes with IC₅₀ values in the low nanomolar range in vitro, possessed favorable pharmacokinetic profiles and showed high efficacy in vivo in several human tumor xenograft models in mice.     

N-(4-(6,7-Disubstituted-quinolin-4-yloxy)-3-fluorophenyl)-2-oxo-3-phenylimidazolidine-1-carboxamides: A novel series of dual c-Met/VEGFR2 receptor tyrosine kinase inhibitors

A series of N-(4-(6,7-disubstituted-quinolin-4-yloxy)-3-fluorophenyl)-2-oxo-3-phenylimidazolidine-1-carboxamides targeting c-Met and VEGFR2 tyrosine kinases was designed and synthesized. The compounds were potent against these two enzymes with IC₅₀ values in the low nanomolar range in vitro, possessed favorable pharmacokinetic profiles and showed high efficacy in vivo in several human tumor xenograft models in mice.     

Biodiversity dynamics in isolated island communities: interaction between natural and human-mediated processes

The flora and fauna of oceanic islands have inspired research since the early scientific explorations. Islands can be considered 'nature's test tubes'- simple systems with multiple replicates. Our research has used the simplicity of island systems to understand ecological community dynamics and to compare the properties of island communities with those in more complex mainland systems. Here, we present three topics: (i) current patterns of biodiversity on isolated islands of the Pacific; (ii) current patterns of disturbance and invasion on islands; and (iii) future trajectories inferred from these patterns. We examine features of islands (in particular, topography and isolation) that have allowed for given levels and distribution of endemicity. The extent to which island communities are impacted by, resist or accommodate disturbance and/or invasions by nonindigenous species appears to be dictated to a large extent by properties of the native communities and how these communities were originally assembled. Accordingly, patterns of disturbance and invasion are very different for high (montane) islands that are extremely isolated compared to those that are nearer to a source of natural migrants. As with all biotas, those on islands are dynamic entities. However, the unique aspect of islands is their isolation, and extreme isolation has largely been lost over the course of the last few centuries due to the development of transportation routes. We argue that such a modified dynamic will affect the future of the biota and the processes that gave rise to the biota. Specifically for isolated habitats, ecological processes will become increasingly more likely to generate biodiversity than evolutionary processes which have been relatively more important in the past. In the short term, island biotas and other similar biotas that occur in montane habitats may fare well as species are often abundant locally in the habitat to which they are indigenous, and may demonstrate considerable resistance and resilience to invasion. However, island biotas - and other biotas that show high local endemism - will likely not fare well in the face of prolonged disturbance. The biotas in these areas generally display a relatively low dispersal capacity; therefore, under conditions of long-term habitat modification, isolated biotas are likely to be swamped by non-natives, which - simply because of random processes and higher propagule pressure - will move more readily into available habitats. Thus, despite the importance of incorporating the evolutionary process into conservation efforts, we must also be careful to evaluate the likely form that the processes will take when the context (specifically, extent of isolation) has been highly modified.     

Biogeography of the fauna of French Polynesia: diversification within and between a series of hot spot archipelagos

The islands of French Polynesia cover an area the size of Europe, though total land area is smaller than Rhode Island. Each hot spot archipelago (Societies, Marquesas, Australs) is chronologically arranged. With the advent of molecular techniques, relatively precise estimations of timing and source of colonization have become feasible. We compile data for the region, first examining colonization (some lineages dispersed from the west, others from the east). Within archipelagos, blackflies (Simulium) provide the best example of adaptive radiation in the Societies, though a similar radiation occurs in weevils (Rhyncogonus). Both lineages indicate that Tahiti hosts the highest diversity. The more remote Marquesas show clear examples of adaptive radiation in birds, arthropods and snails. The Austral Islands, though generally depauperate, host astonishing diversity on the single island of Rapa, while lineages on other islands are generally widespread but with large genetic distances between islands. More recent human colonization has changed the face of Polynesian biogeography. Molecular markers highlight the rapidity of Polynesian human (plus commensal) migrations and the importance of admixture from other populations during the period of prehistoric human voyages. However, recent increase in traffic has brought many new, invasive species to the region, with the future of the indigenous biota uncertain.