Recombinant vacuolar iron transporter family homologue PfVIT from human malaria-causing Plasmodium falciparum is a Fe2+/H+ exchanger

This work was funded in part by a Royal Society Wolfson Research Merit Award (to J.P.D). It was also supported by the Wellcome Trust (grant number WT079272AIA) which funded the MALDI TOF-TOF analyser at the BSRC Mass Spectrometry and Proteomics Facility, University of St Andrews. P.L was supported by a Northern Ireland Department of Employment and Learning (DEL) postgraduate studentship.Vacuolar iron transporters (VITs) are a poorly understood family of integral membrane proteins that can function in iron homeostasis via sequestration of labile Fe2+ into vacuolar compartments. Here we report on the heterologous overexpression and purification of PfVIT, a vacuolar iron transporter homologue from the human malaria-causing parasite Plasmodium falciparum. Use of synthetic, codon-optimised DNA enabled overexpression of functional PfVIT in the inner membrane of Escherichia coli which, in turn, conferred iron tolerance to the bacterial cells. Cells that expressed PfVIT had decreased levels of total cellular iron compared with cells that did not express the protein. Qualitative transport assays performed on inverted vesicles enriched with PfVIT revealed that the transporter catalysed Fe2+/H+ exchange driven by the proton electrochemical gradient. Furthermore, the PfVIT transport function in this system did not require the presence of any Plasmodium-specific factor such as post-translational phosphorylation. PfVIT purified as a monomer and, as measured by intrinsic protein fluorescence quenching, bound Fe2+ in detergent solution with low micromolar affinity. This study of PfVIT provides material for future detailed biochemical, biophysical and structural studies to advance understanding of the vacuolar iron transporter family of membrane proteins from important human pathogens.Publisher PDFPeer reviewe

Aedes aegypti ferritin heavy chain homologue: feeding of iron or blood influences message levels, lengths and subunit abundance

Secreted ferritin in the mosquito, Aedes aegypti, has several subunits that are the products of at least two genes, one encoding a homologue of the vertebrate heavy chain (HCH) and the other the light chain homologue (LCH). Here we report the developmental and organ specific pattern of expression of the ferritin HCH messages and of both subunit types in control sugar-fed mosquitoes, in those exposed to high levels of dietary iron, and after blood feeding. When Northern blots were probed with a HCH cDNA, two bands were observed, representing at least two messages of different sizes that result from the choice of two different polyadenylation sites. Either raising mosquito larvae in an iron-enriched medium, or blood feeding adult female mosquitoes resulted in a marked increase in the HCH message level, particularly of the shorter message. Changes in the amount and length of messages and amount of ferritin subunits were studied over the life span of the mosquito and in different organs of female mosquitoes after blood feeding. The midgut of blood-fed insects is the main site of increased ferritin message synthesis. Ferritin protein levels also increase in midgut, fat body and hemolymph after blood feeding. Ferritin messages and subunits are synthesized in the ovaries and ferritin is found in the eggs. These observations are discussed in terms of translational and transcriptional control of ferritin synthesis and are compared to similar events in the regulation of Drosophila melanogaster ferritin

Lipoprotein biosynthesis in the larvae of the tobacco hornworm, Manduca-sexta

Lipoprotein biosynthesis in larvae of the tobacco hornworm (Manduca sexta) was investigated. By immunoblotting, it was shown that the apoproteins are present in the fat body, but not in the midgut. Fat body incubated in vitro with [35S]methionine secreted labeled apoproteins. However, when the density of the secreted particle was determined, it was found at 1.24-1.28 g/ml instead of 1.15 g/ml, which is the density of the circulating lipoprotein. Lipid analysis of immunoprecipitated lipoprotein secreted by the fat body showed a phospholipid/diacylglycerol ratio of 8.3 rather than 0.9, the ratio found in the circulating lipoprotein. When labeled oleic acid or triolein was fed to larvae, it was found that greater than 98% of the label in the circulating lipoprotein was in diacylglycerol. In studies using animals raised on a fat-free diet, it was shown that the circulating lipoprotein has properties comparable to those of the material secreted in vitro by the fat body and that this diacylglycerol-poor particle can be converted to the normal lipoprotein by feeding a bolus of triolein. These data support the hypothesis that the fat body makes and secretes a "nascent" lipoprotein which contains apoproteins and phospholipid, but is devoid of diacylglycerol. The diacylglycerol is then picked up from the midgut to complete assembly of the mature circulating lipoprotein

Robotic Laser-Adaptive-Optics Imaging of 715 Kepler Exoplanet Candidates using Robo-AO

The Robo-AO Kepler Planetary Candidate Survey is designed to observe every Kepler planet candidate host star with laser adaptive optics imaging to search for blended nearby stars, which may be physically associated companions and/or responsible for transit false positives. In this paper we present the results from the 2012 observing season, searching for stars close to 715 representative Kepler planet candidate hosts. We find 53 companions, 44 of which are new discoveries. We detail the Robo-AO survey data reduction methods including a method of using the large ensemble of target observations as mutual point-spread-function references, along with a new automated companion-detection algorithm designed for large adaptive optics surveys. Our survey is sensitive to objects from 0.15" to 2.5" separation, with contrast ratios up to delta-m~6. We measure an overall nearby-star-probability for Kepler planet candidates of 7.4% +/- 1.0%, and calculate the effects of each detected nearby star on the Kepler-measured planetary radius. We discuss several KOIs of particular interest, including KOI-191 and KOI-1151, which are both multi-planet systems with detected stellar companions whose unusual planetary system architecture might be best explained if they are "coincident multiple" systems, with several transiting planets shared between the two stars. Finally, we detect 2.6-sigma evidence for <15d-period giant planets being 2-3 times more likely be found in wide stellar binaries than smaller close-in planets and all sizes of further-out planets.Comment: Accepted by ApJ. Minor updates & improved statistical analysis; no changes to results. 15 pages, 13 figure

Biochemistry of the sphingolipides. X. Phytoglycolipide, a complex phytosphingosine‐containing lipide from plant seeds

Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/141721/1/aocs0335.pd

Discussion Papers on Irrigation Water Supply Organizations

ii, 61 p. ; 28 cm Irrigation organizations and the reallocation of western water / Rodney T. Smith -- Issues in water conservation / Bruce Driver -- Irrigation districts and water quality / John H. Davidson -- Some thoughts on governing special districts / Tim De Younghttps://scholar.law.colorado.edu/books_reports_studies/1085/thumbnail.jp

Discussion Papers on Irrigation Water Supply Organizations

ii, 61 p. ; 28 cm Irrigation organizations and the reallocation of western water / Rodney T. Smith -- Issues in water conservation / Bruce Driver -- Irrigation districts and water quality / John H. Davidson -- Some thoughts on governing special districts / Tim De Younghttps://scholar.law.colorado.edu/books_reports_studies/1085/thumbnail.jp

Theory of Circle Maps and the Problem of One-Dimensional Optical Resonator with a Periodically Moving Wall

We consider the electromagnetic field in a cavity with a periodically oscillating perfectly reflecting boundary and show that the mathematical theory of circle maps leads to several physical predictions. Notably, well-known results in the theory of circle maps (which we review briefly) imply that there are intervals of parameters where the waves in the cavity get concentrated in wave packets whose energy grows exponentially. Even if these intervals are dense for typical motions of the reflecting boundary, in the complement there is a positive measure set of parameters where the energy remains bounded.Comment: 34 pages LaTeX (revtex) with eps figures, PACS: 02.30.Jr, 42.15.-i, 42.60.Da, 42.65.Y

A New Corpus of Lexical Substitution and Word Blend Errors: Probing the Semantic Structure of Lemma Access Failures

Models of lemma access in language production predict occasional mis-selection of lemmas linked to highly similar concepts (synonyms) and concepts standing in a set-superset relation (subsumatives). It is unclear, however, if such errors occur in spontaneous speech, and if they do, whether humans can detect them given their minimal impact on sentence meaning. This data report examines a large corpus of English spontaneous speech errors and documents a low but non-negligible occurrence of these categories. The existence of synonym and subsumative errors is documented in a larger open access data set that supports a range of new investigations of the semantic structure of lexical substitution and word blend speech errors