Functional Impact of a Cancer-Related Variant in Human Δ\u3csup\u3e1\u3c/sup\u3e‑Pyrroline-5-Carboxylate Reductase 1

Pyrroline-5-carboxylate reductase (PYCR) is a proline biosynthetic enzyme that catalyzes the NAD(P)H-dependent reduction of Δ1-pyrroline-5-carboxylate (P5C) to proline. Humans have three PYCR isoforms, with PYCR1 often upregulated in different types of cancers. Here, we studied the biochemical and structural properties of the Thr171Met variant of PYCR1, which is found in patients with malignant melanoma and lung adenocarcinoma. Although PYCR1 is strongly associated with cancer progression, characterization of a PYCR1 variant in cancer patients has not yet been reported. Thr171 is conserved in all three PYCR isozymes and is located near the P5C substrate binding site. We found that the amino acid replacement does not affect thermostability but has a profound effect on PYCR1 catalytic activity. The kcat of the PYCR1 variant T171M is 100- to 200-fold lower than wild-type PYCR1 when P5C is the variable substrate, and 10- to 25-fold lower when NAD(P)H is varied. A 1.84 Å resolution X-ray crystal structure of T171M reveals that the Met side chain invades the P5C substrate binding site, suggesting that the catalytic defect is due to steric clash preventing P5C from achieving the optimal pose for hydride transfer from NAD(P)H. These results suggest that any impact on PYCR1 function associated with T171M in cancer does not derive from increased catalytic activity

The breakdown of the municipality as caring platform: lessons for co-design and co-learning in the age of platform capitalism

If municipalities were the caring platforms of the 19-20th century sharing economy, how does care manifest in civic structures of the current period? We consider how platforms - from the local initiatives of communities transforming neighbourhoods, to the city, in the form of the local authority - are involved, trusted and/or relied on in the design of shared services and amenities for the public good. We use contrasting cases of interaction between local government and civil society organisations in Sweden and the UK to explore trends in public service provision. We look at how care can manifest between state and citizens and at the roles that co-design and co-learning play in developing contextually sensitive opportunities for caring platforms. In this way, we seek to learn from platforms in transition about the importance of co-learning in political and structural contexts and make recommendations for the co-design of (digital) platforms to care with and for civil society

Deep learning prediction of proton and photon dose distributions for paediatric abdominal tumours

OBJECTIVE: Dose prediction using deep-learning networks prior to radiotherapy might lead to more efficient modality selections. The study goal was to predict proton and photon dose distributions based on the patient-specific anatomy and to assess their clinical usage for paediatric abdominal tumours. MATERIAL &METHODS: Data from 80 patients with neuroblastoma or Wilms' tumour was included. Pencil beam scanning (PBS) (5mm/3%) and volumetric-modulated arc therapy (VMAT) plans (5mm) were robustly optimized on the internal target volume (ITV). Separate 3-dimensional patch-based U-net networks were trained to predict PBS and VMAT dose distributions. Doses, planning-computed tomography images and relevant optimization masks (ITV, vertebra and organs-at-risk) of 60 patients were used for training with a 5-fold cross validation. The networks' performance was evaluated by computing the relative error between planned and predicted dose-volume histogram (DVH) parameters for 20 inference patients. In addition, the organs-at-risk mean dose difference between modalities was calculated using planned and predicted dose distributions (ΔDmean= DVMAT-DPBS). Two radiation oncologists performed a blind PBS/VMAT modality selection based on either planned or predicted ΔDmean. RESULTS: Average DVH differences between planned and predicted dose distributions were ≤|6%|for both modalities. The networks classified the organs-at-risk difference as a gain (ΔDmean>0) with 98% precision. An identical modality selection based on planned compared to predicted ΔDmean was made for 18/20 patients. CONCLUSION: Deep-learning networks for accurate prediction of proton and photon dose distributions for abdominal paediatric tumours were established. These networks allowing fast dose visualization might aid in identifying the optimal radiotherapy technique when experience and/or resources are unavailable

Single quantum dot emission at telecom wavelengths from metamorphic InAs/InGaAs nanostructures grown on GaAs substrates

3 figuras, 3 páginas.We report on the growth by molecular beam epitaxy and the study by atomic force microscopy and photoluminescence of low density metamorphic InAs/InGaAs quantum dots. subcritical InAs coverages allow to obtain 108 cm−2 dot density and metamorphic InxGa1−xAs (x = 0.15,0.30) confining layers result in emission wavelengths at 1.3 μm. We discuss optimal growth parameters and demonstrate single quantum dot emission up to 1350 nm at low temperatures, by distinguishing the main exciton complexes in these nanostructures. Reported results indicate that metamorphic quantum dots could be valuable candidates as single photon sources for long wavelength telecom windows.the financial support of the Generalitat Valenciana and the Spanish Ministry of Science (Project Nos. PROMETEO/2009/074 and TEC2008-06756-C03-03, respectively).Peer reviewe

Potassium and the K\u3csup\u3e+\u3c/sup\u3e/H\u3csup\u3e+\u3c/sup\u3e Exchanger Kha1p Promote Binding of Copper to ApoFet3p Multi-copper Ferroxidase

Acquisition and distribution of metal ions support a number of biological processes. Here we show that respiratory growth of and iron acquisition by the yeast Saccharomyces cerevisiae relies on potassium (K+) compartmentalization to the trans-Golgi network via Kha1p, a K+/H+ exchanger. K+ in the trans-Golgi network facilitates binding of copper to the Fet3p multi-copper ferroxidase. The effect of K+ is not dependent on stable binding with Fet3p or alteration of the characteristics of the secretory pathway. The data suggest that K+ acts as a chemical factor in Fet3p maturation, a role similar to that of cations in folding of nucleic acids. Up-regulation of KHA1 gene in response to iron limitation via iron-specific transcription factors indicates that K+ compartmentalization is linked to cellular iron homeostasis. Our study reveals a novel functional role of K+ in the binding of copper to apoFet3p and identifies a K+/H+ exchanger at the secretory pathway as a new molecular factor associated with iron uptake in yeast

Size dependent carrier thermal escape and transfer in bimodally distributed self assembled InAs/GaAs quantum dots

We have investigated the temperature dependent recombination dynamics in two bimodally distributed InAs self assembled quantum dots samples. A rate equations model has been implemented to investigate the thermally activated carrier escape mechanism which changes from exciton-like to uncorrelated electron and hole pairs as the quantum dot size varies. For the smaller dots, we find a hot exciton thermal escape process. We evaluated the thermal transfer process between quantum dots by the quantum dot density and carrier escape properties of both samples. © 2012 American Institute of Physics.We gratefully acknowledge the financial support of the Generalitat Valenciana, Comunidad Autnoma de Madrid and the Spanish Ministry Projects Nos. PROMETEO/2009/074, S2009ESP-1503 and TEC-2008-06756-C03-03, TEC2011-29120-C05-04/01. One of the authors D. Rivas thanks the Ministry of Science for his FPI fellowship. The AFM characterization has been carried out at CIM, University of Parma, Italy.Peer Reviewe

THERMAL ACTIVATED CARRIER TRANSFER BETWEEN InAs QUANTUM DOTS IN VERY LOW DENSITY SAMPLES

During the last decade, a great effort has been made studying the temperature evolution of QD emission, obtaining good agreements between experimental data and theoretical models. Thermal escape through wetting layer (WL) or by phonon assisted tunneling is usually claimed to describe carrier transfer in monomodal and bimodal QDs distributions. In the present study we have analyzed this phenomenon in two different samples containing a very low density of InAs/GaAs QDs, namely 16.5 and 25 QD/?m2 (Samples I and II, respectively). A detailed experimental study as a function of temperature has been carried out by using ensemble photoluminescence (PL), micro-PL and time resolved PL (TRPL) techniques. In both samples coexist two QD size distributions: (i) a small size one emitting in the region 1.25-1.35 eV (SQD family) and (ii) a large size one emitting in the region 1.05-1.20 eV (LQD family), as shown in Figs. 1.a-b. In sample I the SQD family dominates in intensity and the opposite is observed in Sample II, yet their temperature evolution is similar. An increase of the LQD integrated intensity is observed simultaneously with the decrease of the SDQ band, as observed in Figs. 1.c-d. This behavior is corroborated by the first time by Micro-PL of single QDs (see Fig. 1.e) belonging to both families detected simultaneously. The experiment is performed by using a multimode optical fiber in the detection arm of our confocal microscope and a monomode optical fiber to excite the SQD. A set of balance equations is used to reproduce the measured temperature evolution of the whole PL spectrum by introducing the transfer between SQD towards neighbor LQDs via WL states and the measured TRPL data

Potassium and the K\u3csup\u3e+\u3c/sup\u3e/H\u3csup\u3e+\u3c/sup\u3e Exchanger Kha1p Promote Binding of Copper to ApoFet3p Multi-copper Ferroxidase

Acquisition and distribution of metal ions support a number of biological processes. Here we show that respiratory growth of and iron acquisition by the yeast Saccharomyces cerevisiae relies on potassium (K+) compartmentalization to the trans-Golgi network via Kha1p, a K+/H+ exchanger. K+ in the trans-Golgi network facilitates binding of copper to the Fet3p multi-copper ferroxidase. The effect of K+ is not dependent on stable binding with Fet3p or alteration of the characteristics of the secretory pathway. The data suggest that K+ acts as a chemical factor in Fet3p maturation, a role similar to that of cations in folding of nucleic acids. Up-regulation of KHA1 gene in response to iron limitation via iron-specific transcription factors indicates that K+ compartmentalization is linked to cellular iron homeostasis. Our study reveals a novel functional role of K+ in the binding of copper to apoFet3p and identifies a K+/H+ exchanger at the secretory pathway as a new molecular factor associated with iron uptake in yeast

Cautionary Tale of Using Tris(alkyl)phosphine Reducing Agents with NAD+-Dependent Enzymes

Protein biochemistry protocols typically include disulfide bond reducing agents to guard against unwanted thiol oxidation and protein aggregation. Commonly used disulfide bond reducing agents include dithiothreitol, β-mercaptoethanol, glutathione, and the tris(alkyl)phosphine compounds tris(2-carboxyethyl)phosphine (TCEP) and tris(3-hydroxypropyl)phosphine (THPP). While studying the catalytic activity of the NAD(P)H-dependent enzyme Δ1-pyrroline-5-carboxylate reductase, we unexpectedly observed a rapid non-enzymatic chemical reaction between NAD+ and the reducing agents TCEP and THPP. The product of the reaction exhibits a maximum ultraviolet absorbance peak at 334 nm and forms with an apparent association rate constant of 231–491 M−1 s−1. The reaction is reversible, and nuclear magnetic resonance characterization (1H, 13C, and 31P) of the product revealed a covalent adduct between the phosphorus of the tris(alkyl)phosphine reducing agent and the C4 atom of the nicotinamide ring of NAD+. We also report a 1.45 Å resolution crystal structure of short-chain dehydrogenase/reductase with the NADP+–TCEP reaction product bound in the cofactor binding site, which shows that the adduct can potentially inhibit enzymes. These findings serve to caution researchers when using TCEP or THPP in experimental protocols with NAD(P)+. Because NAD(P)+-dependent oxidoreductases are widespread in nature, our results may be broadly relevant