Molecular Recognition of PTS-1 Cargo Proteins by Pex5p: Implications for Protein Mistargeting in Primary Hyperoxaluria

Peroxisomal biogenesis and function critically depends on the import of cytosolic proteins carrying a PTS1 sequence into this organelle upon interaction with the peroxin Pex5p. Recent structural studies have provided important insights into the molecular recognition of cargo proteins by Pex5p. Peroxisomal import is a key feature in the pathogenesis of primary hyperoxaluria type 1 (PH1), where alanine:glyoxylate aminotransferase (AGT) undergoes mitochondrial mistargeting in about a third of patients. Here, we study the molecular recognition of PTS1 cargo proteins by Pex5p using oligopeptides and AGT variants bearing different natural PTS1 sequences, and employing an array of biophysical, computational and cell biology techniques. Changes in affinity for Pex5p (spanning over 3–4 orders of magnitude) reflect different thermodynamic signatures, but overall bury similar amounts of molecular surface. Structure/energetic analyses provide information on the contribution of ancillary regions and the conformational changes induced in Pex5p and the PTS1 cargo upon complex formation. Pex5p stability in vitro is enhanced upon cargo binding according to their binding affinities. Moreover, we provide evidence that the rational modulation of the AGT: Pex5p binding affinity might be useful tools to investigate mistargeting and misfolding in PH1 by pulling the folding equilibria towards the native and peroxisomal import competent state

Different coordination abilities of 1,7- and 4,7-phenanthroline in the reactions with copper(II) salts: Structural characterization and biological evaluation of the reaction products

The reactions between equimolar amounts of CuX2 (X = NO3− and CF3SO3−) and two aromatic nitrogen-containing heterocycles differing in the position of nitrogen atoms, 1,7- and 4,7-phenanthroline (1,7- and 4,7-phen), were performed in ethanol/methanol at room temperature. When CuX2 salts were mixed with 4,7-phen, two copper(II) complexes, [Cu(NO3)2(4,7-Hphen)2](NO3)2 (1) and [Cu(CF3SO3)(4,7- phen)2(H2O)2]CF3SO3 (2), were formed. On the other hand, in the reaction of CuX2 salts with 1,7-phen, only 1,7-HphenNO3 (3a/b) and 1,7-HphenCF3SO3 (4) were obtained as the final products. The obtained products 1–4 were characterized by spectroscopic and X-ray diffraction techniques. In the copper(II) complexes 1 and 2, the coordination geometry around the Cu(II) ion is distorted octahedral and square pyramidal, respectively. The antimicrobial potential of the copper(II) complexes 1 and 2 and corresponding compounds used for their synthesis were assessed against four different bacterial species and Candida albicans, displaying moderate growth inhibiting activity. The cytotoxic properties of the investigated complexes were also evaluated against the normal human lung fibroblast cell line (MRC-5) indicating moderate, yet more pronounced cytotoxicity than antimicrobial properties

A neuro-fuzzy based combustion sensor for the control of optimal engine combustion efficiency

Modern and advanced control systems for internal combustion engines require accurate feedback information from the combustion chamber. Whereas the in-cylinder pressure sensor provides this information through its close thermodynamic ties with the combustion process, drawbacks in its implementation push research towards other non-intrusive sensing methods. This paper suggests alternative methods of combustion phasing detection relying on measured angular crankshaft speed. Method developed, achieves sensing of angular position of the 50% of mass fraction burned (MFB50) through two steps: calculation of, so called, synthetic torque and its non-linear transformation to a combustion feature estimator through local model. In order to calibrate both parts of this virtual combustion sensor, parameters of a high-fidelity crankshaft dynamic model are identified, and the linear neuro-fuzzy based model is trained with extensive experimentally collected data set. Created virtual MFB50 sensor, demonstrated its performance, on a large test data set comprised of 70% of gathered data

The influence of potassium ion (K+) on digoxin-induced inhibition of porcine cerebral cortex Na+/K+-ATPase

The in vitro influence of potassium ion modulations, in the concentration range 2mM - 500mM, on digoxin-induced inhibition of porcine cerebral cortex Na+/K+-ATPase activity was studied. The response of enzymatic activity in the presence of various K+ concentrations to digoxin was biphasic, thereby, indicating the existence of two Na+/K+-ATPase isoforms, differing in the affinity towards the tested drug. Both isoforms showed higher sensitivity to digoxin in the presence of K+ ions below 20mM in the medium assay. The IC50 values for high/low isoforms 2.77 x 10(-6) M/8.56 x 10(-5) Mand 7.06 x 10(-7) M/1.87 x 10(-5) Mwere obtained in the presence of optimal (20mM) and 2mMK(+), respectively. However, preincubation in the presence of elevated K+ concentration (50 - 500mM) in the medium assay prior to Na+/K+-ATPase exposure to digoxin did not prevent the inhibition, i.e. IC50 values for both isoforms was the same as in the presence of the optimal K+ concentration. On the contrary, addition of 200mMK(+) into the medium assay after 10 minutes exposure of Na+/K+-ATPase to digoxin, showed a time-dependent recovery effect on the inhibited enzymatic activity. Kinetic analysis showed that digoxin inhibited Na+/K+-ATPase by reducing maximum enzymatic velocity (V-max) and K-m, implying an uncompetitive mode of interaction

Molecular Recognition of PTS-1 Cargo Proteins by Pex5p: Implications for Protein Mistargeting in Primary Hyperoxaluria

Peroxisomal biogenesis and function critically depends on the import of cytosolic proteins carrying a PTS1 sequence into this organelle upon interaction with the peroxin Pex5p. Recent structural studies have provided important insights into the molecular recognition of cargo proteins by Pex5p. Peroxisomal import is a key feature in the pathogenesis of primary hyperoxaluria type 1 (PH1), where alanine:glyoxylate aminotransferase (AGT) undergoes mitochondrial mistargeting in about a third of patients. Here, we study the molecular recognition of PTS1 cargo proteins by Pex5p using oligopeptides and AGT variants bearing different natural PTS1 sequences, and employing an array of biophysical, computational and cell biology techniques. Changes in affinity for Pex5p (spanning over 3–4 orders of magnitude) reflect different thermodynamic signatures, but overall bury similar amounts of molecular surface. Structure/energetic analyses provide information on the contribution of ancillary regions and the conformational changes induced in Pex5p and the PTS1 cargo upon complex formation. Pex5p stability in vitro is enhanced upon cargo binding according to their binding affinities. Moreover, we provide evidence that the rational modulation of the AGT: Pex5p binding affinity might be useful tools to investigate mistargeting and misfolding in PH1 by pulling the folding equilibria towards the native and peroxisomal import competent state

Radiation-induced degradation of hydroxyapatite/poly L-lactide composite biomaterial

The effects of gamma irradiation on the structure and properties of hydroxyapatite/poly L-lactide (HAp/PLLA) biomaterial have been investigated. Effects of radiation on microstructure, degradation of polymer part and thermal stability of composite were determined by scanning electronic microscopy (SEM), gel permeation chromatography (GPC) and thermogravimetric analysis (TGA), respectively. Mechanical properties were obtained through mechanical strength and elasticity modulus. Presented results show that properties of HAp/PLLA decay with irradiation dose, but for doses required for sterilization, changes and damaging effects are acceptable. (C) 2006 Elsevier Ltd. All rights reserved