Recent studies indicate that i‐DNA, a four‐stranded cytosine‐rich DNA also known as the i‐motif, is actually formed in vivo; however, a systematic study on sequence effects on stability has been ...missing. Herein, an unprecedented number of different sequences (271) bearing four runs of 3–6 cytosines with different spacer lengths has been tested. While i‐DNA stability is nearly independent on total spacer length, the central spacer plays a special role on stability. Stability also depends on the length of the C‐tracts at both acidic and neutral pHs. This study provides a global picture on i‐DNA stability thanks to the large size of the introduced data set; it reveals unexpected features and allows to conclude that determinants of i‐DNA stability do not mirror those of G‐quadruplexes. Our results illustrate the structural roles of loops and C‐tracts on i‐DNA stability, confirm its formation in cells, and allow establishing rules to predict its stability.
i‐DNA is an emerging non‐canonical DNA secondary structure as an anticancer target and as a basic element in the programmable bionanotechnology. A large number of i‐DNA‐prone sequences were tested to disclose how primary sequences determine the i‐DNA stability both in vitro and in cells.
•pH transition for noninvasive determination of ligand density.•IEX ligands and chelators can be evaluated in a flow-through mode.•pH transition can exhibit flattened or Gaussian-shaped peaks.•The pH ...transition height and length exhibit the same accuracy.•Height is faster and more suitable for small volume or low ligand density.
Stepwise change between low and high salt concentration buffers of the same pH results in pH transition, the length of which was demonstrated to be proportional to the quantity of ion-exchange groups present on the matrix. In this work, we analyzed the effect of the ligand type, density, and buffer concentration on the pH transition shape for typical ion-exchange groups (QA, DEAE, SO3, and COOH) and ligands acting as metal-chelators, such as IDA, TAEA, and EDA. It was demonstrated that pH transition can occur either as a chromatographic or flat-top peak. pH transition peaks were evaluated by their length, height, and peak center parameters. While no parameter can describe the ligand density accurately with a single linear correlation for both peak types, all parameters can be used for the description of one peak type. Peak length and height exhibited the same accuracy, while their sensitivity depended on the pH transition shape: length being more sensitive for the flat-top peaks, while height for the chromatographic peaks. pH height can be obtained faster, at lower elution volume, and seems to be more suitable for the determination of low amounts of ligand, when typically chromatographic peak type pH transitions occur.
The study aims to develop a modification strategy to facilitate uniform catechol-assisted zwitterionization on nitinol alloy for bio-compatibility and fouling resistance. Catechol-functionalized ...polysulfobetaine methacrylate (pSBMA/DA) is synthesized via dopamine-initiated photo-polymerization. Under UV irradiation, semiquinone radicals from dopamine (DA) can be generated, and prevented loss of one electron to intramolecular cyclization and intermolecular dimerization in a solution at pH 2. Pseudo-first-order polymerization kinetics, and relations of apparent rate constant and number average molecular weight with the molar ratio of DA in photopolymerization for pSBMA/DA are unveiled. In a solution at pH 3, PSBMA/DA begins aggregation, kept catechol moieties from premature oxidization, and enabled even deposition on the nitinol substrate. After pH regulation to 8.5, pSBMA/DA extends, and concurrently catechol moieties are activated to interact with the nitinol surface via the formation of bidentate binding. X-ray photoelectron spectroscopy (XPS) analysis revealed that a shorter pSBMA/DA chain with higher catechol content provides more anchoring sites to enhance zwitterionic moieties coverage on substrates. Interestingly, atomic force microscopy (AFM) images revealed a smooth and uniform deposition of pSBMA/DA using the pH-transition method. Strong ionic hydration of pSBMA/DA coating on nitinol surface repels non-specific adsorption of bio-foulants, permitting excellent antifouling properties. Zwitterion-modified nitinol achieved a reduction rate of 99.9% against Escherichia coli and Staphylococcus aureus attachment. In addition, pSBMA/DA exhibits a robust antifouling performance to NIH 3T3 mouse fibroblasts in culture media after incubation for 24 h. Overall, the pSBMA/DA coating via pH transition approach opens up a promising strategy to facilitate uniform surface functionalization for antifouling and coating technology.
Display omitted
•A catechol-assisted polymer was synthesized to functionalize nitinol surface.•Dopamine can be used as a photo-initiator at pH 2 to trigger polymerization.•Catechol moieties within the polymer were in unoxidized form.•Catechol-conjugated polymer can be stably immobilized via pH transition approach.
•Noninvasive quatification of immobilized protein A.•Quantification was demonstrated on polyHIPE monolithic and particulate protein A columns.•Method is based on pH transition profile measurement ...using lactate buffer.•No additional hardware beside HPLC system equipped with pH electrode is required.
The pH transition method, developed for the determination of the ion-exchange group density on chromatographic stationary phase, was used for the quantification of immobilized protein A. Monolithic epoxy polyHIPE and particulate CNBr-Sepharose supports were used for immobilization. A lactate buffer was selected, having a buffer capacity peak approximately 0.5 pH units below the maximum buffer capacity of protein A. The pH transition measurements were performed at pH 4.3, where protein A exhibits maximum buffer capacity, with a lactate buffer concentration of 1 mM for protein A immobilized on polyHIPE monoliths and of 5 mM for protein A immobilized on CNBr-Sepharose. The pH transition height and full width at half maximum for the particulate support and the height for the polyHIPE matrix, showed a linear correlation with the amount of immobilized protein A determined from the absorbance difference before and after immobilization for both supports. The developed method allows a simple, non-invasive on-line determination of immobilized protein A using biological buffers, even for chromatographic columns with an amount of immobilized protein A as low as 0.25 mg. In addition, its sensitivity and duration can be easily adjusted by varying the buffer concentration and pH.
•Pall rings were effectively used for H2S desulfurization under neutral and acid pH.•Oxygen load optimization allowed a minimum S0 production at neutral pH.•Neutral to acid pH transition was ...successfully performed without re-inoculation.•Pyrosequencing (bTEFAP) was successfully used to assess microbial community shifts.•A large simplification of microbial diversity occurred at acidic pH.
Pall rings, a common random packing material, were used in the biotrickling filtration of biogas with high H2S. Assessment of 600d of operation covered the reactor start-up, the operation at neutral pH and the transition from neutral to acid pH. During the start-up period, operational parameters such as the aeration rate and the trickling liquid velocity were optimized. During the steady-state operation at neutral pH, the performance of the random packing material was investigated by reducing the gas contact time at both constant and increasing H2S loads. The random packing material showed similar elimination capacities and removal efficiencies in comparison with previous studies with a structured packing material, indicating that Pall rings are suitable for biogas desulfurization in biotrickling filters. The diversity of Eubacteria and the structure of the community were investigated before and after the pH transition using the bacterial tag-encoded FLX amplicon pyrosequencing. The pH transition to acid pH drastically reduced the microbial diversity and produced a progressive specialization of the sulfur-oxidizing bacteria community without any detrimental effect on the overall desulfurizing capacity of the reactor. During acidic pH operation, a persistent accumulation of elemental sulfur was found.
Recent studies indicate that i‐DNA, a four‐stranded cytosine‐rich DNA also known as the i‐motif, is actually formed in vivo; however, a systematic study on sequence effects on stability has been ...missing. Herein, an unprecedented number of different sequences (271) bearing four runs of 3–6 cytosines with different spacer lengths has been tested. While i‐DNA stability is nearly independent on total spacer length, the central spacer plays a special role on stability. Stability also depends on the length of the C‐tracts at both acidic and neutral pHs. This study provides a global picture on i‐DNA stability thanks to the large size of the introduced data set; it reveals unexpected features and allows to conclude that determinants of i‐DNA stability do not mirror those of G‐quadruplexes. Our results illustrate the structural roles of loops and C‐tracts on i‐DNA stability, confirm its formation in cells, and allow establishing rules to predict its stability.
i‐DNA is an emerging non‐canonical DNA secondary structure as an anticancer target and as a basic element in the programmable bionanotechnology. A large number of i‐DNA‐prone sequences were tested to disclose how primary sequences determine the i‐DNA stability both in vitro and in cells.
The enthalpy change accompanying the reversible acid-induced transition from the native (N) to the molten-globule (MG) state of bovine cytochrome
c was directly evaluated by isothermal acid-titration ...calorimetry (IATC), a new method for evaluating the pH dependence of protein enthalpy. The enthalpy change was 30 kJ/mol at 30 °C, pH 3.54, with 500 mM KCl. The results of the global analysis of the temperature dependence of the excess enthalpy from 20 to 35 °C demonstrated that the N to MG transition is a two-state transition with a small heat capacity change of 1.1 kJ K
−1 mol
−1. The present findings were also indicative of the pH dependence of the enthalpy and the heat capacity of the MG state, −13 kJ mol
−1 pH
−1 and −1.0 kJ K
−1 mol
−1 pH
−1, respectively, at 30 °C within a pH range from 2 to 3.
Pasteurella multocida toxin is a potent mitogen that is believed to act intracellularly. On transverse urea gradient gels at pH 8.0 the toxin displayed one major unfolding transition at 4 M urea. ...However, at pH 6.1 the unfolding transition took place at 3.5 M urea. Circular dichroism spectra also indicated that a structural change took place at acidic pH. In addition it was found that the toxin that had been denatured in 8 M urea refolded in solution with a high recovery of biological activity. These findings are discussed in terms of the likely domain structure of the
P. multocida toxin.
Abstract
Pasteurella multocida toxin is a potent mitogen that is believed to act intracellularly. On transverse urea gradient gels at pH 8.0 the toxin displayed one major unfolding transition at 4 M ...urea. However, at pH 6.1 the unfolding transition took place at 3.5 M urea. Circular dichroism spectra also indicated that a structural change took place at acidic pH. In addition it was found that the toxin that had been denatured in 8 M urea refolded in solution with a high recovery of biological activity. These findings are discussed in terms of the likely domain structure of the P. multocida toxin.
PDF
