Voltage-dependent structural changes of the membrane-bound anion channel hVDAC1 probed by SEIRA and electrochemical impedance spectroscopy

Dieser Beitrag ist mit Zustimmung des Rechteinhabers aufgrund einer (DFG geförderten) Allianz- bzw. Nationallizenz frei zugänglich.This publication is with permission of the rights owner freely accessible due to an Alliance licence and a national licence (funded by the DFG, German Research Foundation) respectively.The voltage-dependent anion channel (VDAC) is a transmembrane protein that regulates the transfer of metabolites between the cytosol and the mitochondrium. Opening and partial closing of the channel is known to be driven by the transmembrane potential viaa mechanism that is not fully understood. In this work, we employed a spectroelectrochemical approach to probe the voltage-induced molecular structure changes of human VDAC1 (hVDAC1) embedded in a tethered bilayer lipid membrane on a nanostructured Au electrode. The model membrane consisted of a mixed self-assembled monolayer of 6-mercaptohexanol and (cholesterylpolyethylenoxy)thiol, followed by the deposition of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine vesicles including hVDAC1. The stepwise assembly of the model membrane and the incorporation of hVDAC1 were monitored by surface enhanced infrared absorption and electrochemical impedance spectroscopy. Difference spectra allowed for identifying the spectral changes which may be associated with the transition from the open to the “closed” states by shifting the potential above or below the transmembrane potential determined to beca.0.0 Vvs.the open circuit potential. These spectral changes were interpreted on the basis of the orientation- and distance-dependent IR enhancement and indicate alterations of the inclination angle of the β-strands as crucial molecular events, reflecting an expansion or contraction of the β-barrel pore. These protein structural changes that do not confirm nor exclude the reorientation of the α-helix are either directly induced by the electric field or a consequence of a potential-dependent repulsion or attraction of the bilayer.DFG, EXC 314, Unifying Concepts in CatalysisDFG, SFB 803, Funktionalität kontrolliert durch Organisation in und zwischen Membrane

Thermodynamics and kinetics of reduction and species conversion at a hydrophobic surface for mitochondrial cytochromes c and their cardiolipin adducts

Cytochrome c(cytc) and its adduct with cardiolipin (CL) were immobilized on a hydrophobic SAM-coated electrode surface yielding a construct which mimics the environment experienced by the complex at the inner mitochondrial membrane where it plays a role in cell apoptosis. Under these conditions, both species undergo an equilibrium between a six-coordinated His/His-ligated and a five-coordinated His/- ligated forms stable in the oxidized and in the reduced state, respectively. The thermodynamics of the oxidation-state dependent species conversion were determined by temperature-dependent diffusionless voltammetry experiments. CL binding stabilizes the immobilized reduced His/- ligated form of cytc which was found previously to catalytically reduce dioxygen. Here, this adduct is also found to show pseudoperoxidase activity, catalysing reduction of hydrogen peroxide. These effects would impart CL with an additional role in the cytc-mediated peroxidation leading to programmed cell death. Moreover, Immobilized cytc exchanges electrons more slowly upon CL binding possibly due to changes in solvent reorganization effects at the protein-SAM interface

Mapping local electric fields in proteins at biomimetic interfaces

Dieser Beitrag ist mit Zustimmung des Rechteinhabers aufgrund einer (DFG geförderten) Allianz- bzw. Nationallizenz frei zugänglich.This publication is with permission of the rights owner freely accessible due to an Alliance licence and a national licence (funded by the DFG, German Research Foundation) respectively.We present a novel approach for determining the strength of the electric field experienced by proteins immobilised on membrane models. It is based on the vibrational Stark effect of a nitrile label introduced at different positions on engineered proteins and monitored by surface enhanced infrared absorption spectroscopy

Active-site structure, binding and redox activity of the heme–thiolate enzyme CYP2D6 immobilized on coated Ag electrodes: a surface-enhanced resonance Raman scattering study

Surface-enhance resonance Raman scattering spectra of the heme–thiolate enzyme cytochrome P450 2D6 (CYP2D6) adsorbed on Ag electrodes coated with 11-mercaptoundecanoic acid (MUA) were obtained in various experimental conditions. An analysis of these spectra, and a comparison between them and the RR spectra of CYP2D6 in solution, indicated that the enzyme’s active site retained its nature of six-coordinated low-spin heme upon immobilization. Moreover, the spectral changes detected in the presence of dextromethorphan (a CYP2D6 substrate) and imidazole (an exogenous heme axial ligand) indicated that the immobilized enzyme also preserved its ability to reversibly bind a substrate and form a heme–imidazole complex. The reversibility of these processes could be easily verified by flowing alternately solutions of the various compounds and the buffer through a home-built spectroelectrochemical flow cell which contained a sample of immobilized protein, without the need to disassemble the cell between consecutive spectral data acquisitions. Despite immobilized CYP2D6 being effectively reduced by a sodium dithionite solution, electrochemical reduction via the Ag electrode was not able to completely reduce the enzyme, and led to its extensive inactivation. This behavior indicated that although the enzyme’s ability to exchange electrons is not altered by immobilization per se, MUA-coated electrodes are not suited to perform direct electrochemistry of CYP2D6

Colorectal cancer after bariatric surgery (Cric-Abs 2020): Sicob (Italian society of obesity surgery) endorsed national survey

Background The published colorectal cancer (CRC) outcomes after bariatric surgery (BS) are conflicting, with some anecdotal studies reporting increased risks. The present nationwide survey CRIC-ABS 2020 (Colo-Rectal Cancer Incidence-After Bariatric Surgery-2020), endorsed by the Italian Society of Obesity Surgery (SICOB), aims to report its incidence in Italy after BS, comparing the two commonest laparoscopic procedures-Sleeve Gastrectomy (SG) and Roux-en-Y gastric bypass (GBP). Methods Two online questionnaires-first having 11 questions on SG/GBP frequency with a follow-up of 5-10 years, and the second containing 15 questions on CRC incidence and management, were administered to 53 referral bariatric, high volume centers. A standardized incidence ratio (SIR-a ratio of the observed number of cases to the expected number) with 95% confidence intervals (CI) was calculated along with CRC incidence risk computation for baseline characteristics. Results Data for 20,571 patients from 34 (63%) centers between 2010 and 2015 were collected, of which 14,431 had SG (70%) and 6140 GBP (30%). 22 patients (0.10%, mean age = 53 +/- 12 years, 13 males), SG: 12 and GBP: 10, developed CRC after 4.3 +/- 2.3 years. Overall incidence was higher among males for both groups (SG: 0.15% vs 0.05%; GBP: 0.35% vs 0.09%) and the GBP cohort having slightly older patients. The right colon was most affected (n = 13) and SIR categorized/sex had fewer values < 1, except for GBP males (SIR = 1.07). Conclusion Low CRC incidence after BS at 10 years (0.10%), and no difference between procedures was seen, suggesting that BS does not trigger the neoplasm development

Zinc-Alpha 2-Glycoprotein Gene Expression in Adipose Tissue Is Related with Insulin Resistance and Lipolytic Genes in Morbidly Obese Patients

10.1371/journal.pone.003326

Infected pancreatic necrosis: outcomes and clinical predictors of mortality. A post hoc analysis of the MANCTRA-1 international study

: The identification of high-risk patients in the early stages of infected pancreatic necrosis (IPN) is critical, because it could help the clinicians to adopt more effective management strategies. We conducted a post hoc analysis of the MANCTRA-1 international study to assess the association between clinical risk factors and mortality among adult patients with IPN. Univariable and multivariable logistic regression models were used to identify prognostic factors of mortality. We identified 247 consecutive patients with IPN hospitalised between January 2019 and December 2020. History of uncontrolled arterial hypertension (p = 0.032; 95% CI 1.135-15.882; aOR 4.245), qSOFA (p = 0.005; 95% CI 1.359-5.879; aOR 2.828), renal failure (p = 0.022; 95% CI 1.138-5.442; aOR 2.489), and haemodynamic failure (p = 0.018; 95% CI 1.184-5.978; aOR 2.661), were identified as independent predictors of mortality in IPN patients. Cholangitis (p = 0.003; 95% CI 1.598-9.930; aOR 3.983), abdominal compartment syndrome (p = 0.032; 95% CI 1.090-6.967; aOR 2.735), and gastrointestinal/intra-abdominal bleeding (p = 0.009; 95% CI 1.286-5.712; aOR 2.710) were independently associated with the risk of mortality. Upfront open surgical necrosectomy was strongly associated with the risk of mortality (p < 0.001; 95% CI 1.912-7.442; aOR 3.772), whereas endoscopic drainage of pancreatic necrosis (p = 0.018; 95% CI 0.138-0.834; aOR 0.339) and enteral nutrition (p = 0.003; 95% CI 0.143-0.716; aOR 0.320) were found as protective factors. Organ failure, acute cholangitis, and upfront open surgical necrosectomy were the most significant predictors of mortality. Our study confirmed that, even in a subgroup of particularly ill patients such as those with IPN, upfront open surgery should be avoided as much as possible. Study protocol registered in ClinicalTrials.Gov (I.D. Number NCT04747990)

Relationships between xylem anatomy, root hydraulic conductivity, leaf/root ratio and transpiration in citrus trees on different rootstocks

The aim of the study was to determine the extent in which leaf and whole plant transpiration (Tp) were influenced by root hydraulic conductance (K(r)), leaf to root ratio and leaf mass. Also, the relationships between the anatomic characteristics of roots and K(r) were investigated. To this end, 9-month-old seedlings of the citrus rootstocks Cleopatra mandarin (CM), Poncirus trifoliata (PT), and their hybrids Forner-Alcaide no 5 (FA-5) and Forner-Alcaide no 13 (FA-13) and 15-month-old trees of Valencia orange budded on these four rootstocks were tested. The hybrid FA-13 and PT had higher values of K(r) and leaf transpiration rates (E) than FA-5 and CM. There was a positive curvilinear correlation between E and K(r). Furthermore, E levels in the different types of plants decreased with increased leaf/root (L/R) ratios. Pruning of the roots and defoliation confirmed that transpiration rates were strongly influenced by the L/R ratio. However, variations in E because of differences in L/R ratios were less pronounced in trees budded on FA-13 and PT than on the other two rootstocks. In addition, there was a positive correlation between Tp and leaf biomass, although differences between rootstocks may be attributed to differences in K(r). The average lumen diameter of xylem vessels was greater in rootstocks with high K(r). Size of epidermal and hypodermal cells of fibrous roots may also restrict K(r)