Structural and biochemical characterization of NarE, an iron-containing ADP-ribosyltransferase from Neisseria meningitidis

NarE is a 16 kDa protein identified from Neisseria meningitidis, one of the bacterial pathogens responsible for meningitis. NarE belongs to the family of ADP-ribosyltransferases (ADPRT) and catalyzes the transfer of ADP-ribose moieties to arginine residues in target protein acceptors. Many pathogenic bacteria utilize ADP-ribosylating toxins to modify and alter essential functions of eukaryotic cells. NarE is further the first ADPRT which could be shown to bind iron through a Fe-S center, which is crucial for the catalytic activity. Here we present the NMR solution structure of NarE, which shows structural homology to other ADPRTs. Using NMR titration experiments we could identify from Chemical Shift Perturbation data both the NAD binding site, which is in perfect agreement with a consensus sequence analysis between different ADPRTs, as well as the iron coordination site, which consists of 2 cysteines and 2 histidines. This atypical iron coordination is also capable to bind zinc. These results could be fortified by site-directed mutagenesis of the catalytic region, which identified two functionally crucial residues. We could further identify a main interaction region of NarE with antibodies using two complementary methods based on antibody immobilization, proteolytic digestion, and mass spectrometry. This study combines structural and functional features of NarE providing for the first time a characterization of an iron-dependent ADPRT.     

One‐to‐one registration of en‐face optical coherence tomography attenuation coefficients with histology of a prostatectomy specimen

Optical coherence tomography (OCT), enables high‐resolution 3D imaging of the morphology of light scattering tissues. From the OCT signal, parameters can be extracted and related to tissue structures. One of the quantitative parameters is the attenuation coefficient; the rate at which the intensity of detected light decays in depth. To couple the quantitative parameters with the histology one‐to‐one registration is needed. The primary aim of this study is to validate a registration method of quantitative OCT parameters to histological tissue outcome through one‐to‐one registration of OCT with histology. We matched OCT images of unstained fixated prostate tissue slices with corresponding histology slides, wherein different histologic types were demarcated. Attenuation coefficients were determined by a supervised automated exponential fit (corrected for point spread function and sensitivity roll‐off related signal losses) over a depth of 0.32 mm starting from 0.10 mm below the automatically detected tissue edge. Finally, the attenuation coefficients corresponding to the different tissue types of the prostate were compared. From the attenuation coefficients, we produced the squared relative residue and goodness‐of‐fit metric R². This article explains the method to perform supervised automated quantitative analysis of OCT data, and the one‐to‐one registration of OCT extracted quantitative data with histopathological outcomes.      

Arginine-Specific Mono ADP-Ribosylation In Vitro of Antimicrobial Peptides by ADP-Ribosylating Toxins

Among the several toxins used by pathogenic bacteria to target eukaryotic host cells, proteins that exert ADP-ribosylation activity represent a large and studied family of dangerous and potentially lethal toxins. These proteins alter cell physiology catalyzing the transfer of the ADP-ribose unit from NAD to cellular proteins involved in key metabolic pathways. In the present study, we tested the capability of four of these toxins, to ADP-ribosylate α- and β- defensins. Cholera toxin (CT) from Vibrio cholerae and heat labile enterotoxin (LT) from Escherichia coli both modified the human α-defensin (HNP-1) and β- defensin-1 (HBD1), as efficiently as the mammalian mono-ADP-ribosyltransferase-1. Pseudomonas aeruginosa exoenzyme S was inactive on both HNP-1 and HBD1. Neisseria meningitidis NarE poorly recognized HNP-1 as a substrate but it was completely inactive on HBD1. On the other hand, HNP-1 strongly influenced NarE inhibiting its transferase activity while enhancing auto-ADP-ribosylation. We conclude that only some arginine-specific ADP-ribosylating toxins recognize defensins as substrates in vitro. Modifications that alter the biological activities of antimicrobial peptides may be relevant for the innate immune response. In particular, ADP-ribosylation of antimicrobial peptides may represent a novel escape mechanism adopted by pathogens to facilitate colonization of host tissues.     

Increasing Strawberry Fruit Sensorial and Nutritional Quality Using Wild and Cultivated Germplasm

Background

Increasing antioxidant levels in fruit through breeding is an important option to support higher antioxidant intake particularly when fruit consumption is low. Indeed, if nutritional components are also combined with a high standard of sensorial fruit quality, the perspective for consumer health can be further improved by encouraging more fruit consumption. Wild species are valued by strawberry breeders as sources of novel traits, especially for pest resistance and abiotic stress tolerance. Furthermore, previous investigations have shown improvements in fruit nutritional quality in breeding material that originated from Fragaria virginiana ssp. glauca (FVG) inter-species crosses. Recently, commercial varieties of strawberries have also shown interesting variability in fruit nutritional quality.

Results

Strawberry fruit sensorial and nutritional qualities generated by Fragaria inter-species and intra-species crosses were evaluated on 78 offspring derived from 8 families: two that originated from F. × ananassa intra-species crossing; three from back-crossing of F1– FVG × F. × ananassa; and three from back-crossing of BC1– FVG × F. × ananassa. The genetic variability from the three types of cross combinations was analyzed by calculation of the correlations among the fruit sensorial and nutritional parameters. The results obtained show that two subsequent back-crossing generations from an inter-species crossing combination with F. virginiana ssp. glauca provides useful improvement of the fruit nutritional and sensorial qualities that is combined with agronomic standards that are close to those requested at the commercial level. Improvements of these traits can also be achieved by programming F. × ananassa intra-species crosses and producing progeny with productivity traits more similar to those of the commercial cultivars.

Conclusions

The two types of combination programs (inter-species back-crosses, and intra-species crosses) can be used to improve strawberry nutritional quality.     

Numerical simulation of the dynamics of a bileaflet prosthetic heart valve using a fluid-structure interaction approach

The main purpose of this study is to reproduce in silico the dynamics of a bileaflet mechanical heart valve (MHV; St Jude Hemodynamic Plus, 27mm characteristic size) by means of a fully implicit fluid-structure interaction (FSI) method, and experimentally validate the results using an ultrafast cinematographic technique. The computational model was constructed to realistically reproduce the boundary condition (72 beats per minute (bpm), cardiac output 4.5l/min) and the geometry of the experimental setup, including the valve housing and the hinge configuration. The simulation was carried out coupling a commercial computational fluid dynamics (CFD) package based on finite-volume method with user-defined code for solving the structural domain, and exploiting the parallel performance of the whole numerical setup. Outputs are leaflets excursion from opening to closure and the fluid dynamics through the valve. Results put in evidence a favorable comparison between the computed and the experimental data: the model captures the main features of the leaflet motion during the systole. The use of parallel computing drastically limited the computational costs, showing a linear scaling on 16 processors (despite the massive use of user-defined subroutines to manage the FSI process). The favorable agreement obtained between in vitro and in silico results of the leaflet displacements confirms the consistency of the numerical method used, and candidates the application of FSI models to become a major tool to optimize the MHV design and eventually provides useful information to surgeons.     

Enabling Fluorine-Free Lithium-Ion Capacitors and Lithium-Ion Batteries for High-Temperature Applications by the Implementation of Lithium Bis(oxalato)Borate and Ethyl Isopropyl Sulfone as Electrolyte

A novel fluorine-free electrolyte comprising a solution of lithium bis(oxalato)borate in ethyl isopropyl sulfone is presented. It is characterized by its safety and non-toxic properties, along with the capability to effectively suppress the anodic dissolution of aluminum. Successful high-temperature application of this electrolyte in combination with various capacitor- and battery-like electrode materials is shown. Further utilization in a lithium-ion capacitor and a lithium-ion battery is demonstrated. To the best of the knowledge, the lithium-ion capacitor presented in this work represents the first entirely fluorine-free device suitable for high-temperature applications. When operating at 60 °C, this device delivers a maximum energy output of 169 Wh kg⁻¹_AM at a power of 200 W kg⁻¹_AM and even 80 Wh kg¹_AM at 10 kW kg^-1_AM, along with the ability to retain 80% of its initial capacitance after 3500 cycles at 5 A g⁻¹. As such, this novel electrolyte is a promising alternative to conventional fluorine-containing configurations since its performance is capable to match or even surpass that of most similar laboratory-scale LICs.     

Impact of warming and drought on carbon balance related to wood formation in black spruce

Background and Aims

Wood formation in trees represents a carbon sink that can be modified in the case of stress. The way carbon metabolism constrains growth during stress periods (high temperature and water deficit) is now under debate. In this study, the amounts of non-structural carbohydrates (NSCs) for xylogenesis in black spruce, Picea mariana, saplings were assessed under high temperature and drought in order to determine the role of sugar mobilization for osmotic purposes and its consequences for secondary growth.

Methods

Four-year-old saplings of black spruce in a greenhouse were subjected to different thermal conditions with respect to the outside air temperature (T0) in 2010 (2 and 5 °C higher than T0) and 2011 (6 °C warmer than T0 during the day or night) with a dry period of about 1 month in June of each year. Wood formation together with starch, NSCs and leaf parameters (water potential and photosynthesis) were monitored from May to September.

Key Results

With the exception of raffinose, the amounts of soluble sugars were not modified in the cambium even if gas exchange and photosynthesis were greatly reduced during drought. Raffinose increased more than pinitol under a pre-dawn water potential of less than –1 Mpa, presumably because this compound is better suited than polyol for replacing water and capturing free radicals, and its degradation into simple sugar is easier. Warming decreased the starch storage in the xylem as well the available hexose pool in the cambium and the xylem, probably because of an increase in respiration.

Conclusions

Radial stem growth was reduced during drought due to the mobilization of NSCs for osmotic purposes and due to the lack of cell turgor. Thus plant water status during wood formation can influence the NSCs available for growth in the cambium and xylem.