Geminate rebinding in R-state hemoglobin: kinetic and computational evidence for multiple hydrophobic pockets

Biphasic geminate rebinding of CO to myoglobin upon flash photolysis has been associated to ligand distribution in hydrophobic cavities, structurally detected by time-resolved crystallography, xenon occupancy, and molecular simulations. We show that the time course of CO rebinding to human hemoglobin also exhibits a biphasic geminate rebinding when the protein is entrapped in wet nanoporous silica gel. A simple branched kinetic scheme, involving the bound state A, the primary docking site C, and a secondary binding site B was used to calculate the microscopic rates and the time-dependent population of the intermediate species. The activation enthalpies of the associated transitions were determined in the absence and presence of 80% glycerol. Potential hydrophobic docking cavities within the alpha and beta chains of hemoglobin were identified by computational modeling using xenon as a probe. A hydrophobic pocket on the distal side of the heme, corresponding to Xe4 in Mb, and a nearby site that does not have a correspondence in Mb were detected. Neither potential xenon sites on the proximal side nor a migration channel from the distal to proximal site was located. The small enthalpic barriers between states B and C are in very good agreement with the location of the xenon sites on the distal side. Furthermore, the connection between the two xenon sites is relatively open, explaining why the decreased mobility of the protein with viscosity only slightly perturbs the energetics of ligand migration between the two sites.     

Prediction of intracellular storage polymers using quantitative image analysis in enhanced biological phosphorus removal systems

The present study focuses on predicting the concentration of intracellular storage polymers in enhanced biological phosphorus removal (EBPR) systems. For that purpose, quantitative image analysis techniques were developed for determining the intracellular concentrations of PHA (PHB and PHV) with Nile blue and glycogen with aniline blue staining. Partial least squares (PLS) were used to predict the standard analytical values of these polymers by the proposed methodology. Identification of the aerobic and anaerobic stages proved to be crucial for improving the assessment of PHA, PHB and PHV intracellular concentrations. Current Nile blue based methodology can be seen as a feasible starting point for further enhancement. Glycogen detection based on the developed aniline blue staining methodology combined with the image analysis data proved to be a promising technique, toward the elimination of the need for analytical off-line measurements.     

Magnetic properties of a polyfluorene derivative containing complexed neodymium ions

Poly[9,9′‐dihexylfluorene‐2,7‐diyl)‐6,6″‐(2,2′:6′,2″‐terpyridine)] (LaPPS75) and its complexes with neodymium were synthesized and characterized. Magnetic measurements showed that the noncomplexed polymer presented a ferromagnetic contribution due to the formation of π stacking, and that in absence of those, the ferromagnetic behavior is suppressed. The pristine polymer, the complexed one and a low‐molecular‐weight model compound with the same structure of the complexed site in the parent polymer were studied. The observed behavior found is presented and discussed, the most important finding was that when a conjugated chain is used as a host for the metallic ion, an amplification of four times for the magnetization is achieved, using the same metallic content for complexed polymer and model compound for comparison. © 2019 Wiley Periodicals, Inc. J. Polym. Sci., Part B: Polym. Phys. 2019 , 57, 304–311     

Low‐Level Laser Therapy Restores the Oxidative Stress Balance in Acute Lung Injury Induced by Gut Ischemia and Reperfusion

It remains unknown if the oxidative stress can be regulated by low‐level laser therapy (LLLT) in lung inflammation induced by intestinal reperfusion (i‐I/R). A study was developed in which rats were irradiated (660 nm, 30 mW, 5.4 J) on the skin over the bronchus and euthanized 2 h after the initial of intestinal reperfusion. Lung edema and bronchoalveolar lavage fluid neutrophils were measured by the Evans blue extravasation and myeloperoxidase (MPO) activity respectively. Lung histology was used for analyzing the injury score. Reactive oxygen species (ROS) was measured by fluorescence. Both expression intercellular adhesion molecule 1 (ICAM‐1) and peroxisome proliferator‐activated receptor‐y (PPARy) were measured by RT‐PCR. The lung immunohistochemical localization of ICAM‐1 was visualized as a brown stain. Both lung HSP70 and glutathione protein were evaluated by ELISA. LLLT reduced neatly the edema, neutrophils influx, MPO activity and ICAM‐1 mRNA expression. LLLT also reduced the ROS formation and oppositely increased GSH concentration in lung from i‐I/R groups. Both HSP70 and PPARy expression also were elevated after laser irradiation. Results indicate that laser effect in attenuating the acute lung inflammation is driven to restore the balance between the pro‐ and antioxidants mediators rising of PPARy expression and consequently the HSP70 production.     

Substituent Effects on EI-MS Fragmentation Patterns of 5-Allyloxy-1-aryl-tetrazoles and 4-Allyl-1-aryl-tetrazole-5-ones; Correlation with UV-Induced Fragmentation Channels

1,4- and 1,5-disubstituted tetrazoles possess enriched structures and versatile chemistry, representing a challenge for chemists. In the present work, we unravel the fragmentation patterns of a chemically diverse range of 5-allyloxy-1-aryl-tetrazoles and 4-allyl-1-aryl-tetrazolole-5-ones when subjected to electron impact mass spectrometry (EI-MS) and investigate the correlation with the UV-induced fragmentation channels of the matrix-isolated tetrazole derivatives. Our results indicate that the fragmentation pathways of the selected tetrazoles in EI-MS are highly influenced by the electronic effects induced by substitution. Multiple pathways can be envisaged to explain the mechanisms of fragmentation, frequently awarding common final species, namely arylisocyanate, arylazide, arylnitrene, isocyanic acid and hydrogen azide radical cations, as well as allyl/aryl cations. The identified fragments are consistent with those found in previous investigations concerning the photochemical stability of the same class of molecules. This parallelism showcases a similarity in the behaviour of tetrazoles under EI-MS and UV-irradiation in the inert environment of cryogenic matrices of noble gases, providing efficient tools for reactivity predictions, whether for analytical ends or more in-depth studies. Theoretical calculations provide complementary information to articulate predictions of resulting products.     

Controlling the Surface Functionalization of Ultrasmall Gold Nanoparticles by Sequence-Defined Macromolecules

Ultrasmall gold nanoparticles (diameter about 2 nm) were surface-functionalized with cysteine-carrying precision macromolecules. These consisted of sequence-defined oligo(amidoamine)s (OAAs) with either two or six cysteine molecules for binding to the gold surface and either with or without a PEG chain (3400 Da). They were characterized by ¹H NMR spectroscopy, ¹H NMR diffusion-ordered spectroscopy (DOSY), small-angle X-ray scattering (SAXS), and high-resolution transmission electron microscopy. The number of precision macromolecules per nanoparticle was determined after fluorescent labeling by UV spectroscopy and also by quantitative ¹H NMR spectroscopy. Each nanoparticle carried between 40 and 100 OAA ligands, depending on the number of cysteine units per OAA. The footprint of each ligand was about 0.074 nm² per cysteine molecule. OAAs are well suited to stabilize ultrasmall gold nanoparticles by selective surface conjugation and can be used to selectively cover their surface. The presence of the PEG chain considerably increased the hydrodynamic diameter of both dissolved macromolecules and macromolecule-conjugated gold nanoparticles.