Structural Basis for Carbapenemase Activity of the OXA-23 β-Lactamase from Acinetobacter baumannii

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Growth of vertically aligned carbon nanotubes on carbon fiber: thermal and electrochemical treatments

Composite electrodes of vertically aligned carbon nanotubes (VACNT) were synthesized on carbon fiber (CF) substrate by pyrolysis of camphor/ferrocene using a SiO₂ interlayer as a barrier against metal diffusion into the substrate. Two treatments were used to remove iron from CF/VACNT structure: thermal annealing at high temperature under inert atmosphere and electrochemical oxidation in H₂SO₄ solution. The composites were characterized by scanning electron microscopy and Raman scattering spectroscopy. Besides, the electrochemical behavior of CF/VACNT was analyzed by cyclic voltammetry and charge/discharge tests. CF/VACNT composite submitted to the electrochemical oxidation showed the best electrochemical performance, with high specific capacitance, which makes it very attractive as electrode for supercapacitors.     

Performance of hydrophobic interaction ligands for human membrane‐bound catechol‐O‐methyltransferase purification

Despite of membrane catechol‐O‐methyltransferase (MBCOMT, EC 2.1.1.6) physiological importance on catecholamines’ O‐methylation, no studies allowed their total isolation. Therefore, for the first time, we compare the performance of three hydrophobic adsorbents (butyl‐, epoxy‐, and octyl‐Sepharose) in purification of recombinant human COMT (hMBCOMT) from crude Brevibacillus choshinensis cell lysates to develop a sustainable chromatographic process. Hydrophobic matrices were evaluated in terms of selectivity and hMBCOMT's binding and elution conditions. Results show that hMBCOMT's adsorption was promoted on octyl and butyl at ≤375 mM NaH₂PO_4, while on epoxy higher concentrations (>850 mM) were required. Additionally, hMBCOMT's elution was promoted on epoxy, butyl, and octyl using respectively 0.1–0.5, 0.25–1, and 1% of Triton X‐100. On butyl media, a stepwise strategy using 375 and 0 mM NaH₂PO₄, followed by three elution steps at 0.25, 0.7 and 1% Triton X‐100, allowed selective hMBCOMT isolation. In conclusion, significant amounts of MBCOMT were purified with high selectivity on a single chromatography procedure, despite its elution occurs on multiple peaks. Although successful applications of hydrophobic interaction chromatography in purification of membrane proteins are uncommon, we proved that traditional hydrophobic matrices can open a promising unexplored field to fulfill specific requirements for kinetic and pharmacological trials.     

Ionization by pH and Anionic Surfactant Binding Gives the Same Thickening Effects of Crosslinked Polyacrylic Acid Derivatives

Physical properties of aqueous solutions of hydrophobically modified crosslinked polyacrylic acids change quite extensively as the polymer is charged up. A study is carried out concerning the similarities between two polymer ionization processes, that is, by pH increment and anionic surfactant addition. The two processes charge the polymer by distinctly different mechanisms. At sufficiently high pH the carboxylic groups of the polymer are virtually all ionized and the polymer is, therefore, fully charged. The effective repulsion among the charged groups due to the entropy of the counterions promotes an increased stiffness as well as an expansion of the polymer particles. We investigate here how the ionization and swelling will be if, instead of high pH, the polymer is at low pH conditions but associated to ionic surfactants. Surfactants associate to the polymer both in a noncooperative way by the binding of individual surfactant molecules and in a cooperative way as micelles since the polymer promotes surfactant self-assembly. This binding leads to a highly charged polymer-surfactant complex and leads to an osmotic swelling as well. The swelling and the gelation were monitored by rheology and dynamic light scattering, of polymer solutions by varying the pHs and adding ionic surfactants at low pH. The results show that ionization by surfactants and by pH lead to approximately the same gelation degree, as can be seen by similar viscosity values. Both processes result in dramatic viscosity increases, up to 8 orders of magnitude. More hydrophobic surfactants, with longer alkyl chain, are shown to be more efficient as enhancers of swelling and gelation. The network that is formed at high pH or at sufficiently high concentration of surfactant can be weakened or even disrupted if monovalent or divalent salts are added, demonstrating the role of counterion entropy.     

A Universal DNA Aptamer that Recognizes Spike Proteins of Diverse SARS-CoV-2 Variants of Concern

We report on a unique DNA aptamer, denoted MSA52, that displays universally high affinity for the spike proteins of wildtype SARS-CoV-2 as well as the Alpha, Beta, Gamma, Epsilon, Kappa, Delta and Omicron variants. Using an aptamer pool produced from round 13 of selection against the S1 domain of the wildtype spike protein, we carried out one-round SELEX experiments using five different trimeric spike proteins from variants, followed by high-throughput sequencing and sequence alignment analysis of aptamers that formed complexes with all proteins. A previously unidentified aptamer, MSA52, showed K_d values ranging from 2 to 10 nM for all variant spike proteins, and also bound similarly to variants not present in the reselection experiments. This aptamer also recognized pseudotyped lentiviruses (PL) expressing eight different spike proteins of SARS-CoV-2 with K_d values between 20 and 50 pM, and was integrated into a simple colorimetric assay for detection of multiple PL variants. This discovery provides evidence that aptamers can be generated with high affinity to multiple variants of a single protein, including emerging variants, making it well-suited for molecular recognition of rapidly evolving targets such as those found in SARS-CoV-2.     

Inter-network magnetic interactions in GdMexMn1−xO3 perovskites (Me=transition metal)

The gadolinium-based manganite GdMnO₃ of perovskite structure has been partially substituted at the manganese site by transition metal elements Me like Cu, Ni and Co, leading to a general formula GdMe_xMn_1−xO₃, in which different magnetic entities (e.g., Gd³⁺, Cu²⁺, Ni²⁺, Co²⁺, Co³⁺, Mn³⁺, Mn⁴⁺) can coexist, depending on charge equilibrium conditions. For divalent cations such as Cu²⁺ and Ni²⁺, the solid solution extends from x  (Me)=0–0.5, with O-type orthorhombic symmetry (a<c/√2<b)$\left(a<c/\surd 2<b\right)$. When the substituting element is cobalt, the solid solution extends over the whole range [0?x  ?1], changing from O′-type symmetry (c/√2<a<b)$\left(c/\surd 2<a<b\right)$ to O-type for x>0.5. In this latter case, the synthesis is performed under oxygen flow, which allows the cobalt ion to take a 3+ oxidation state.     

Turning Tryptophanase into Odor‐Generating Biosensors

An odor‐based sensor system that exploits the metabolic enzyme tryptophanase (TPase) as the key component is reported. This enzyme is able to convert an odorless substrate like S‐methyl‐L ‐cysteine or L ‐tryptophan into the odorous products methyl mercaptan or indole. To make a biosensor, TPase was biotinylated so that it could be coupled with a molecular recognition element, such as an antibody, to develop an ELISA‐like assay. This method was used for the detection of an antibody present in nM concentrations by the human nose. TPase can also be combined with the enzyme pyridoxal kinase (PKase) for use in a coupled assay to detect adenosine 5′‐triphosphate (ATP). When ATP is present in the low μM concentration range, the coupled enzymatic system generates an odor that is easily detectable by the human nose. Biotinylated TPase can be combined with various biotin‐labeled molecular recognition elements, thereby enabling a broad range of applications for this odor‐based reporting system.     

Pullulan Encapsulation of Labile Biomolecules to Give Stable Bioassay Tablets

A simple and inexpensive method is reported for the long‐term stabilization of enzymes and other unstable reagents in premeasured quantities in water‐soluble tablets (cast, not compressed) made with pullulan, a nonionic polysaccharide that forms an oxygen impermeable solid upon drying. The pullulan tablets dissolve in aqueous solutions in seconds, thereby facilitating the easy execution of bioassays at remote sites with no need for special reagent handling and liquid pipetting. This approach is modular in nature, thus allowing the creation of individual tablets for enzymes and their substrates. Proof‐of‐principle demonstrations include a Taq polymerase tablet for DNA amplification through PCR and a pesticide assay kit consisting of separate tablets for acetylcholinesterase and its chromogenic substrate, indoxyl acetate, both of which are highly unstable. The encapsulated reagents remain stable at room temperature for months, thus enabling the room‐temperature shipping and storage of bioassay components.     

A comparative multi-reference configuration interaction study of the low-lying states of two thione isomers of thiophenol

Multi-reference configuration interaction, MR-CI (including extensivity corrections, named +Q), calculations were performed on the S₀–S₃ states of cyclohexa-2,4-diene-1-thione (thione 24 ) and cyclohexa-2,5-diene-1-thione (thione 25 ), which are thione isomers of thiophenol. Several types of uncontracted MR-CIS and MR-CISD wavefunctions were employed, comprising MR-CI expansions as large as ~365 × 10⁶ configuration state functions. The nature of the studied excited states was characterized. Vertical excitation energies (ΔE) and oscillator strengths (f) were computed. The most intense transitions (S₀ → S₂ for 24 and S₀ → S₃ for 25 ) did not change with the wavefunction, although a variation as large as ~1 eV was obtained for the S₃ state of 24 , at the highest (MR-CI+Q) level. On the other hand, ΔE changed at most by ~0.56 eV for 25 as the wavefunction changes, at the same level. The S₁ state of both thiones was found to have nπ* character and is in the visible region. For 24 , S₂ and S₃ are ππ* and nπ* states, respectively, while for 25 the reverse order is obtained. S₂ and S₃ are in the range ~3.5 to 5.2 eV, again at the highest level. It is the first time that the excited states of the title molecules are studied. The computed results agree with the experimental onset of photoreactions of thiones 24 and 25 found by Reva et al (Phys. Chem. Chem. Phys., 2015 , 17, 4888).     

Evaluation of the cytotoxicity on breast cancer cell of extracts and compounds isolated from Hyptis pectinata (L.) poit

Abstract

Hyptis pectinata is a herb popularly used in Brazil for the treatment of inflammations, pain, bacterial infections and cancer. In the present study, inflorescences (MPIn), leaves (MPL), branches (MPB), root (MPR) extracts and three compounds isolated from MPIn were assayed against breast tumor cell lines. The structures of the three compounds (pectinolide J, hyptolide and pectinolide E) were determined by means of spectroscopic analysis. Pectinolide J was isolated for the first time. The MPIn, MPL and MPR exhibited specific antiproliferative activity on tumor cell lines when compared to normal cell lines with IC₅₀ of 52.01?±?0.64, 45.91?±?0.02?μg/mL and 82.84?±?0.03?μg/mL, respectively. Although the isolated substances did not present good antiproliferative activity, when the three were associated, a greater biological effect was observed, suggesting a synergistic effect. Hyptolide (5.6?±?0.4?μg/mL) showed IC₅₀ sufficiently low to be considered as a drug prototype.