Fort CnoX: Protecting Bacterial Proteins From Misfolding and Oxidative Damage

How proteins fold and are protected from stress-induced aggregation is a long-standing mystery and a crucial question in biology. Here, we present the current knowledge on the chaperedoxin CnoX, a novel type of protein folding factor that combines holdase chaperone activity with a redox protective function. Focusing on Escherichia coli CnoX, we explain the essential role played by this protein under HOCl (bleach) stress, discussing how it protects its substrates from both aggregation and irreversible oxidation, which could otherwise interfere with refolding. Finally, we highlight the unique ability of CnoX, apparently conserved during evolution, to cooperate with the GroEL/ES folding machinery.

The Providence Mutation (βK82D) in Human Hemoglobin Substantially Reduces βCysteine 93 Oxidation and Oxidative Stress in Endothelial Cells

The highly toxic oxidative transformation of hemoglobin (Hb) to the ferryl state (HbFe4+) is known to occur in both in vitro and in vivo settings. We recently constructed oxidatively stable human Hbs, based on the Hb Providence (βK82D) mutation in sickle cell Hb (βE6V/βK82D) and in a recombinant crosslinked Hb (rHb0.1/βK82D). Using High Resolution Accurate Mass (HRAM) mass spectrometry, we first quantified the degree of irreversible oxidation of βCys93 in these proteins, induced by hydrogen peroxide (H2O2), and compared it to their respective controls (HbA and HbS). Both Hbs containing the βK82D mutation showed considerably less cysteic acid formation, a byproduct of cysteine irreversible oxidation. Next, we performed a novel study aimed at exploring the impact of introducing βK82D containing Hbs on vascular endothelial redox homeostasis and energy metabolism. Incubation of the mutants carrying βK82D with endothelial cells resulted in altered bioenergetic function, by improving basal cellular glycolysis and glycolytic capacity. Treatment of cells with Hb variants containing βK82D resulted in lower heme oxygenase-1 and ferritin expressions, compared to native Hbs. We conclude that the presence of βK82D confers oxidative stability to Hb and adds significant resistance to oxidative toxicity. Therefore, we propose that βK82D is a potential gene-editing target in the treatment of sickle cell disease and in the design of safe and effective oxygen therapeutics.

Engineering biocompatible TeSex nano-alloys as a versatile theranostic nanoplatform

Photothermal nanotheranostics, especially in the near infrared II (NIR-II) region, exhibits a great potential in precision and personalized medicine, owing to high tissue penetration of NIR-II light. NIR-II-photothermal nanoplatforms with high biocompatibility as well as high photothermal effect are urgently needed but rarely reported so far. Te nanomaterials possess high absorbance to NIR-II light but also exhibit high cytotoxicity, impeding their biomedical applications. In this work, the controllable incorporation of biocompatible Se into the lattice of Te nanostructures is proposed to intrinsically tune their inherent cytotoxicity and enhance their biocompatibility, developing TeSex nano-alloys as a new kind of theranostic nanoplatform. We have uncovered that the cytotoxicity of Te nanomaterials primarily derives from irreversible oxidation stress and intracellular imbalance of organization and energy, and can be eliminated by incorporating a moderate proportion of Se (x = 0.43). We have also discovered that the as-prepared TeSex nano-alloys have extraordinarily high NIR-II-photothermal conversion efficiency (77.2%), 64Cu coordination and computed tomography contrast capabilities, enabling high-efficacy multimodal photothermal/photoacoustic/positron emission tomography/computed tomography imaging-guided NIR-II-photothermal therapy of cancer. The proposed nano-alloying strategy provides a new route to improve the biocompatibility of biomedical nanoplatforms and endow them with versatile theranostic functions.

Syntheses, theoretical studies, and crystal structures of [Ni(II)SSRRL](PF6)2 and [Ni(II)SRSRL](Cl)(PF6) that contains anagostic interactions

The syntheses of two square planar nickel complexes containing the condensation and subsequently reduced products obtained by reacting [Ni(en)3](BF4)2 and acetone are reported. The complexes 5,5,7,12,12,14-hexamethyl-1(S),4(S),8(R),11(R)-tetraazacyclotetradecane-nickel(II)[PF6]2 and 5,5,7,12,12,14-hexamethyl-1(S),4(R),8(S),11(R)-tetraazacyclotetradecane-nickel(II)[Cl][PF6] labelled as [Ni(II)SSRRL](PF6)2 and [Ni(II)SRSRL](Cl)(PF6), respectively, were found to have slightly different solubilities that allowed for their purification. The complexes were characterized by FTIR, 1H NMR, and UV–vis spectra. Redox potentials, determined by cyclic voltammetry, established that [Ni(II)SSRRL](PF6)2 exhibits a reversible oxidation (E1/2(ox) = 0.85 V) and reduction (E1/2(red) = −1.59 V), whereas [Ni(II)SRSRL](Cl)(PF6) displays an irreversible oxidation (Epa(ox) = 1.37 V) and reversible reduction (E1/2(red) = −1.62 V) relative to the ferrocene couple at 0.0 V. Single crystal X-ray determinations established that one of the compounds, [Ni(II)SSRRL](PF6)2, contained two [Formula: see text] anions, whereas the other compound, [Ni(II)SRSRL](Cl)(PF6), contained one Cl− and one [Formula: see text] anion. In the solid state, compound [Ni(II)SSRRL](PF6)2 was held together by H-bonds between H atoms on the Ni containing dication and F atoms in the [Formula: see text] anion. Compound [Ni(II)SRSRL](Cl)(PF6) crystallized in the form of dimers held together by interactions between H atoms attached to N atoms on adjacent cations binding to two Cl− anions in the middle with these dimers held together by further H-bonding to interstitial [Formula: see text] anions. Complex [Ni(II)SRSRL](Cl)(PF6) was found to contain anagostic interactions on the bases of NMR (downfield shift in C–H protons) and structural data (2.3 < d(H-Ni) < 2.9 Å), as well as theoretical calculations.

The redox status of cysteine thiol residues of apolipoprotein E impacts on its lipid interactions

Redox-mediated modulation of cysteine (Cys) thiols has roles in various pathophysiological functions. We recently found that formation of disulfide-linked complexes of apolipoprotein (apo) E3 prevented apoE3 from irreversible oxidation. In this report, the influence of modification of Cys thiols in apoE2 and apoE3 on interactions with lipids was investigated. The apoE redox status was examined by a band-shift assay using a maleimide compound, and interactions with lipids were evaluated by a kinetic assay using dimyristoyl-sn-glycero-3-phosphocholine (DMPC) and non-denaturing polyacrylamide gel electrophoresis. A reduction in DMPC clearance activity of apoE2 and apoE3 but not apoE4 was observed. Although hydrogen peroxide-induced oxidation decreased the clearance activity of the isoforms, apoE2 showed the greatest residual activity. Both Cys thiol masking and dimerization decreased the activity of apoE2 and apoE3 but not apoE4. In contrast, apoAII preincubation markedly increased the activity (apoE2 > apoE3 > apoE4), in accordance with the formation of apoE-AII and apoAII-E2-AII complexes. ApoAII preincubation also reduced the particle size of apoE-DMPC liposome complexes, especially for apoE2. Redox-mediated modification of Cys thiols of apoE2 or apoE3, especially disulfide bond formation with apoAII, affects lipid metabolism and consequently may be responsible for the diverse isoform specificity of apoE.

Control of protein function through oxidation and reduction of persulfidated states

Irreversible oxidation of Cys residues to sulfinic/sulfonic forms typically impairs protein function. We found that persulfidation (CysSSH) protects Cys from irreversible oxidative loss of function by the formation of CysSSO1-3H derivatives that can subsequently be reduced back to native thiols. Reductive reactivation of oxidized persulfides by the thioredoxin system was demonstrated in albumin, Prx2, and PTP1B. In cells, this mechanism protects and regulates key proteins of signaling pathways, including Prx2, PTEN, PTP1B, HSP90, and KEAP1. Using quantitative mass spectrometry, we show that (i) CysSSH and CysSSO3H species are abundant in mouse liver and enzymatically regulated by the glutathione and thioredoxin systems and (ii) deletion of the thioredoxin-related protein TRP14 in mice altered CysSSH levels on a subset of proteins, predicting a role for TRP14 in persulfide signaling. Furthermore, selenium supplementation, polysulfide treatment, or knockdown of TRP14 mediated cellular responses to EGF, suggesting a role for TrxR1/TRP14-regulated oxidative persulfidation in growth factor responsiveness.

An Ru-Substituted Tris(ethynyl)methyl Cation: Synthesis, Properties, and Structure of [{Cp(dppe)Ru(C≡C)}3C]PF6·C6H6

The dark blue complex [{Cp(dppe)Ru(C≡C)}3C]PF6 1 (Cp=cyclopentadienyl, dppe=1,2-bis(diphenylphosphino)ethane) was obtained in 46% yield by treatment of Ru(C≡CH)(dppe)Cp with CuCl/TMEDA (tetramethylethanediamine), followed by KOH and [NH4]PF6 in acetone; it was accompanied by known complexes {Cp(dppe)Ru}C≡CC≡C{Ru(dppe)Cp} 2 (22%) and yellow [1,3-{Cp(dppe)Ru}2C4H3]PF6 3 (2.6%). The structure of the cationic fragment of 1 in its benzene solvate consists of a central planar C attached to three C≡CRu(dppe)Cp fragments. The cation of 3 consists of a cyclobuten-1,3-diyl group bearing two Ru(dppe)Cp groups. The 13C NMR resonance of the central C in 1 is found at δ 66.11. The cyclic voltammogram of 1 contains three irreversible oxidation waves at +0.87, +0.79, and +0.25V, together with a reversible reduction wave at −1.38V (versus FeCp2/[FeCp2]+).

Generation of Retinaldehyde for Retinoic Acid Biosynthesis

The concentration of all-trans-retinoic acid, the bioactive derivative of vitamin A, is critically important for the optimal performance of numerous physiological processes. Either too little or too much of retinoic acid in developing or adult tissues is equally harmful. All-trans-retinoic acid is produced by the irreversible oxidation of all-trans-retinaldehyde. Thus, the concentration of retinaldehyde as the immediate precursor of retinoic acid has to be tightly controlled. However, the enzymes that produce all-trans-retinaldehyde for retinoic acid biosynthesis and the mechanisms responsible for the control of retinaldehyde levels have not yet been fully defined. The goal of this review is to summarize the current state of knowledge regarding the identities of physiologically relevant retinol dehydrogenases, their enzymatic properties, and tissue distribution, and to discuss potential mechanisms for the regulation of the flux from retinol to retinaldehyde.