Case report: Precise NGS and combined bevacizumab promote durable response in ALK-positive lung adenocarcinoma with multiple-line ALK-TKI resistance.

Liquid biopsies including pleural fluid or plasma are commonly applied for patients with advanced non-small cell lung cancer (NSCLC) and pleural effusion (PE) to guide the treatment. ALK-TKIs are the first options for patients with ALK-positive mutations and combining ALK-TKIs with angiogenic agents may improve survival. We report here one case with ALK-positive lung adenocarcinoma in which the patient achieved a prolonged progression-free survival (PFS) of 97 months after undergoing precise pleural effusion NGS and receiving combined bevacizumab treatment following multiple-line ALK-TKI resistance.

Updated roles of the gut microbiota in exploring shrimp etiology, polymicrobial pathogens, and disease incidence.

Litopenaeus vannamei is the most extensively cultured shrimp species globally, recognized for its scale, production, and economic value. However, its aquaculture is plagued by frequent disease outbreaks, resulting in rapid and massive mortality. etiological research often lags behind the emergence of new diseases, leaving the causal agents of some shrimp diseases unidentified and leading to nomenclature based on symptomatic presentations, especially in cases involving co- and polymicrobial pathogens. Comprehensive data on shrimp disease statuses remain limited. In this review, we summarize current knowledge on shrimp diseases and their effects on the gut microbiome. Furthermore, we also propose a workflow integrating primary colonizers, "driver" taxa in gut networks from healthy to diseased states, disease-discriminatory taxa, and virulence genes to identify potential polymicrobial pathogens. We examine both abiotic and biotic factors (e.g., external and internal sources and specific-disease effects) that influence shrimp gut microbiota, with an emphasis on the "holobiome" concept and common features of gut microbiota response to diverse diseases. After excluding the effects of confounding factors, we provide a diagnosis model for quantitatively predicting shrimp disease incidence using disease common-discriminatory taxa, irrespective of the causal agents. Due to the conservation of functional genes used in designing specific primers, we propose a practical strategy applying qPCR-assayed abundances of disease common-discriminatory functional genes. This review updates the roles of the gut microbiota in exploring shrimp etiology, polymicrobial pathogens, and disease incidence, offering a refined perspective for advancing shrimp aquaculture health management.

Observation of oxygenated intermediates in functional mimics of aminophenol dioxygenase.

Aminophenol dioxygenases (APDO) are mononuclear nonheme iron enzymes that utilize dioxygen (O2) to catalyze the conversion of o-aminophenols to 2-picolinic acid derivatives in metabolic pathways. This study describes the synthesis and O2 reactivity of two synthetic models of substrate-bound APDO: [FeII(TpMe2)(tBu2APH)] (1) and [FeII(TpMe2)(tBuAPH)] (2), where TpMe2 = hydrotris(3,5-dimethylpyrazole-1-yl)borate, tBu2APH = 4,6-di-tert-butyl-2-aminophenolate, and tBuAPH2 = 4-tert-butyl-2-aminophenolate. Both Fe(II) complexes behave as functional APDO mimics, as exposure to O2 results in oxidative CC bond cleavage of the o-aminophenolate ligand. The ring-cleaved products undergo spontaneous cyclization to give substituted 2-picolinic acids, as verified by 1H NMR spectroscopy, mass spectrometry, and X-ray crystallography. Reaction of the APDO models with O2 at low temperature reveals multiple intermediates, which were probed with UV-vis absorption, electron paramagnetic resonance (EPR), Mössbauer (MB), and resonance Raman (rRaman) spectroscopies. The most stable intermediate at -70 °C in THF exhibits multiple isotopically-sensitive features in rRaman samples prepared with 16O2 and 18O2, confirming incorporation of O2-derived atom(s) into its molecular structure. Insights into the geometric structures, electronic properties, and spectroscopic features of the observed intermediates were obtained from density functional theory (DFT) calculations. Although functional APDO models have been previously reported, this is the first time that an oxygenated ligand-based radical has been detected and spectroscopically characterized in the ring-cleaving mechanism of a relevant synthetic system.

Theoretical comparison of fructose with methylglucoside for the production of formate and levulinate catalyzed by Brønsted acids in a methanol solution.

For the conversion of fructose/methylglucoside (MG) into both methyl formate (MF) and methyl levulinate (MLev), the C-source of formate [HCOO]- remains unclear at the molecular level. Herein, reaction mechanisms catalyzed by [CH3OH2]+ in a methanol solution were theoretically investigated at the PBE0/6-311++G(d,p) level. For the conversion of fructose into MF and MLev, the formate [HCOO]- comes from the C1-atom of fructose, in which the rate-determining step lies in the reaction of 5-hydroxymethylfurfural (HMF) with CH3OH to yield MF and MLev. The reaction of fructose with CH3OH kinetically tends to generate HMF intermediates rather than yield (MF + MLev). When MG is dissolved in a methanol solution, its O2, O3, and O4 atoms are closer to the first layer of the solvent than O1, O5, and O6 atoms. For the dehydration of MG with methanol into MF and MLev, the formate [HCOO]- stems from the dominant C1- and secondary C3-atoms of MG. Kinetically, MG is ready to yield (MF + MLev), whereas fructose can induce the reaction to remain at the HMF intermediate, inhibiting the further conversion of HMF with CH3OH into MF and MLev. If MG isomerizes into fructose, the reaction will be more preferable for yielding HMF rather than (MF + MLev).

Temperature-dependent iron motion in extremophile rubredoxins - no need for 'corresponding states'.

Extremophile organisms are known that can metabolize at temperatures down to - 25 °C (psychrophiles) and up to 122 °C (hyperthermophiles). Understanding viability under extreme conditions is relevant for human health, biotechnological applications, and our search for life elsewhere in the universe. Information about the stability and dynamics of proteins under environmental extremes is an important factor in this regard. Here we compare the dynamics of small Fe-S proteins - rubredoxins - from psychrophilic and hyperthermophilic microorganisms, using three different nuclear techniques as well as molecular dynamics calculations to quantify motion at the Fe site. The theory of 'corresponding states' posits that homologous proteins from different extremophiles have comparable flexibilities at the optimum growth temperatures of their respective organisms. Although 'corresponding states' would predict greater flexibility for rubredoxins that operate at low temperatures, we find that from 4 to 300 K, the dynamics of the Fe sites in these homologous proteins are essentially equivalent.

Catalytic Activity of the Archetype from Group 4 of the FTR-like Ferredoxin:Thioredoxin Reductase Family Is Regulated by Unique S = 7/2 and S = 1/2 [4Fe-4S] Clusters.

Thioredoxin reductases (TrxR) activate thioredoxins (Trx) that regulate the activity of diverse target proteins essential to prokaryotic and eukaryotic life. However, very little is understood of TrxR/Trx systems and redox control in methanogenic microbes from the domain Archaea (methanogens), for which genomes are abundant with annotations for ferredoxin:thioredoxin reductases [Fdx/thioredoxin reductase (FTR)] from group 4 of the widespread FTR-like family. Only two from the FTR-like family are characterized: the plant-type FTR from group 1 and FDR from group 6. Herein, the group 4 archetype (AFTR) from Methanosarcina acetivorans was characterized to advance understanding of the family and TrxR/Trx systems in methanogens. The modeled structure of AFTR, together with EPR and Mössbauer spectroscopies, supports a catalytic mechanism similar to plant-type FTR and FDR, albeit with important exceptions. EPR spectroscopy of reduced AFTR identified a transient [4Fe-4S]1+ cluster exhibiting a mixture of S = 7/2 and typical S = 1/2 signals, although rare for proteins containing [4Fe-4S] clusters, it is most likely the on-pathway intermediate in the disulfide reduction. Furthermore, an active site histidine equivalent to residues essential for the activity of plant-type FTR and FDR was found dispensable for AFTR. Finally, a unique thioredoxin system was reconstituted from AFTR, ferredoxin, and Trx2 from M. acetivorans, for which specialized target proteins were identified that are essential for growth and other diverse metabolisms.

Mechanism of CO2 in promoting the hydrogenation of levulinic acid to γ-valerolactone catalyzed by RuCl3 in aqueous solution.

A Ru-containing complex shows good catalytic performance toward the hydrogenation of levulinic acid (LA) to γ-valerolactone (GVL) with the assistance of organic base ligands (OBLs) and CO2. Herein, we report the competitive mechanisms for the hydrogenation of LA to GVL, 4-oxopentanal (OT), and 2-methyltetrahydro-2,5-furandiol (MFD) with HCOOH or H2 as the H source catalyzed by RuCl3 in aqueous solution at the M06/def2-TZVP, 6-311++G(d,p) theoretical level. Kinetically, the hydrodehydration of LA to GVL is predominant, with OT and MFD as side products. With HCOOH as the H source, initially, the OBL (triethylamine, pyridine, or triphenylphosphine) is responsible for capturing H+ from HCOOH, leading to HCOO- and [HL]+. Next, the Ru3+ site is in charge of sieving H- from HCOO-, yielding [RuH]2+ hydride and CO2. Alternatively, with H2 as the H source, the OBL stimulates the heterolysis of H-H bond with the aid of Ru3+ active species, producing [RuH]2+ and [HL]+. Toward the [RuH]2+ formation, H2 as the H source exhibits higher activity than HCOOH as the H source in the presence of an OBL. Thereafter, H- in [RuH]2+ gets transferred to the unsaturated C site of ketone carbonyl in LA. Afterwards, the Ru3+ active species is capable of cleaving the C-OH bond in 4-hydroxyvaleric acid, yielding [RuOH]2+ hydroxide and GVL. Subsequently, CO2 promotes Ru-OH bond cleavage in [RuOH]2+, forming HCO3- and regenerating the Ru3+-active species owing to its Lewis acidity. Lastly, between the resultant HCO3- and [HL]+, a neutralization reaction occurs, generating H2O, CO2, and OBLs. Thus, the present study provides insights into the promotive roles of additives such as CO2 and OBLs in Ru-catalyzed hydrogenation.

NMR and Mössbauer Studies Reveal a Temperature-Dependent Switch from S = 1 to 2 in a Nonheme Oxoiron(IV) Complex with Faster C-H Bond Cleavage Rates.

S = 2 FeIV═O centers generated in the active sites of nonheme iron oxygenases cleave substrate C-H bonds at rates significantly faster than most known synthetic FeIV═O complexes. Unlike the majority of the latter, which are S = 1 complexes, [FeIV(O)(tris(2-quinolylmethyl)amine)(MeCN)]2+ (3) is a rare example of a synthetic S = 2 FeIV═O complex that cleaves C-H bonds 1000-fold faster than the related [FeIV(O)(tris(pyridyl-2-methyl)amine)(MeCN)]2+ complex (0). To rationalize this significant difference, a systematic comparison of properties has been carried out on 0 and 3 as well as related complexes 1 and 2 with mixed pyridine (Py)/quinoline (Q) ligation. Interestingly, 2 with a 2-Q-1-Py donor combination cleaves C-H bonds at 233 K with rates approaching those of 3, even though Mössbauer analysis reveals 2 to be S = 1 at 4 K. At 233 K however, 2 becomes S = 2, as shown by its 1H NMR spectrum. These results demonstrate a unique temperature-dependent spin-state transition from triplet to quintet in oxoiron(IV) chemistry that gives rise to the high C-H bond cleaving reactivity observed for 2.

Reconstitution of the Final Steps in the Biosynthesis of Valanimycin Reveals the Origin of Its Characteristic Azoxy Moiety.

Valanimycin is an azoxy-containing natural product isolated from the fermentation broth of Streptomyces viridifaciens MG456-hF10. While the biosynthesis of valanimycin has been partially characterized, how the azoxy group is constructed remains obscure. Herein, the membrane protein VlmO and the putative hydrazine synthetase ForJ from the formycin biosynthetic pathway are demonstrated to catalyze N-N bond formation converting O-(l-seryl)-isobutyl hydroxylamine into N-(isobutylamino)-l-serine. Subsequent installation of the azoxy group is shown to be catalyzed by the non-heme diiron enzyme VlmB in a reaction in which the N-N single bond in the VlmO/ForJ product is oxidized by four electrons to yield the azoxy group. The catalytic cycle of VlmB appears to begin with a resting µ-oxo diferric complex in VlmB, as supported by Mössbauer spectroscopy. This study also identifies N-(isobutylamino)-d-serine as an alternative substrate for VlmB leading to two azoxy regioisomers. The reactions catalyzed by the kinase VlmJ and the lyase VlmK during the final steps of valanimycin biosynthesis are established as well. The biosynthesis of valanimycin was thus fully reconstituted in vitro using the enzymes VlmO/ForJ, VlmB, VlmJ and VlmK. Importantly, the VlmB-catalyzed reaction represents the first example of enzyme-catalyzed azoxy formation and is expected to proceed by an atypical mechanism.

Electroacupuncture at Sensitized Acupoints Relieves Somatic Referred Pain in Colitis Rats by Inhibiting Sympathetic-Sensory Coupling to Interfere with 5-HT Signaling Pathway.

OBJECTIVE: To investigate whether electroacupuncture (EA) at sensitized acupoints could reduce sympathetic-sensory coupling (SSC) and neurogenic inflammatory response by interfering with 5-hydroxytryptamine (5-HT)ergic neural pathways to relieve colitis and somatic referred pain, and explore the underlying mechanisms. METHODS: Rats were treated with 5% dextran sodium sulfate (DSS) solution for 7 days to establish a colitis model. Twelve rats were randomly divided into the control and model groups according to a random number table (n=6). According to the "Research on Rat Acupoint Atlas", sensitized acupoints and non-sensitized acupoints were determined. Rats were randomly divided into the control, model, Zusanli-EA (ST 36), Dachangshu-EA (BL 25), and Xinshu (BL 15) groups (n=6), as well as the control, model, EA, and EA + GR113808 (a 5-HT inhibitor) groups (n=6). The rats in the control group received no treatment. Acupuncture was administered on 2 days after modeling using the stimulation pavameters: 1 mA, 2 Hz, for 30 min, with sparse and dense waves, for 14 consecutive days. GR113808 was injected into the tail vein at 5 mg/kg before EA for 10 min for 7 consecutive days. Mechanical sensitivity was assessed with von Frey filaments. Body weight and disease activity index (DAI) scores of rats were determined. Hematoxylin and eosin staining was performed to observe colon histopathology. SSC was analyzed by immunofluorescence staining. Immunohistochemical staining was performed to detect 5-HT and substance P (SP) expressions. The calcitonin gene-related peptide (CGRP) in skin tissue and tyrosine hydroxylase (TH) protein levels in DRG were detected by Western blot. The levels of hyaluronic acid (HA), bradykinin (BK), prostaglandin I2 (PGI2) in skin tissue, 5-HT, tryptophan hydroxylase 1 (TPH1), serotonin transporters (SERT), 5-HT 3 receptor (5-HT3R), and 5-HT 4 receptor (5-HT4R) in colon tissue were measured by enzyme-linked immunosorbent assay (ELISA). RESULTS: BL 25 and ST 36 acupoints were determined as sensitized acupoints, and BL 15 acupoint was used as a non-sensitized acupoint. EA at sensitized acupoints improved the DAI score, increased mechanical withdrawal thresholds, and alleviated colonic pathological damage of rats. EA at sensitized acupoints reduced SSC structures and decreased TH and CGRP expression levels (P<0.05). Furthermore, EA at sensitized acupoints reduced BK, PGI2, 5-HT, 5-HT3R and TPH1 levels, and increased HA, 5-HT4R and SERT levels in colitis rats (P<0.05). GR113808 treatment diminished the protective effect of EA at sensitized acupoints in colitis rats (P<0.05). CONCLUSION: EA at sensitized acupoints alleviated DSS-induced somatic referred pain in colitis rats by interfering with 5-HTergic neural pathway, and reducing SSC inflammatory response.

Formation of a Reactive [Mn(III)-O-Ce(IV)] Species and its Facile Equilibrium with Related Mn(IV)(OX) (X = Sc or H) Complexes.

Lewis acid-bound high valent Mn-oxo species are of great importance due to their relevance to photosystem II. Here, we report the synthesis of a unique [(BnTPEN)Mn(III)-O-Ce(IV)(NO3 )4 ]+ adduct (2) by the reaction of (BnTPEN)Mn(II) (1) with 4 eq. ceric ammonium nitrate. 2 has been characterized using UV/Vis, NMR, resonance Raman spectroscopy, as well as by mass spectrometry. Treatment of 2 with Sc(III)(OTf)3 results in the formation of (BnTPEN)Mn(IV)-O-Sc(III) (3), while HClO4 addition to 2 forms (BnTPEN)Mn(IV)-OH (4), reverting to 2 upon Ce(III)(NO3 )3 addition. 2 can also be prepared by the oxidation of 1 eq. Ce(III)(NO3 )3 with [(BnTPEN)Mn(IV)=O]2+ (5). In addition, the EPR spectroscopy revealed the elegant temperature-dependent equilibria between 2 and Mn(IV) species. The binding of redox-active Ce(IV) boosts electron transfer efficiency of 2 towards ferrocenes. Remarkably, the newly characterized Mn(III)-O-Ce(IV) species can carry out O-atom and H-atom transfer reactions.

Polyiodo Azole-Based Metal-Organic Framework Energetic Biocidal Material for Synergetic Sterilization Applications.

Biological hazards caused by bacteria, viruses, and toxins have become a major survival and development issue facing the international community. However, the traditional method of disinfection and sterilization is helpless in dealing with viruses that spread quickly and are highly infectious. Metal-organic framework (MOF) biocidal materials hold promise as superior alternatives to traditional sterilization materials because of their stable framework structures and unique properties. Now, we demonstrate for the first time the synthesis of a MOF (TIBT-Cu) containing Cu metal centers and tetraiodo-4,4'-bi-1,2,4-triazole as the main ligand. This novel MOF biocidal material has good thermal stability (Td = 278 °C), excellent mechanical sensitivity, and a high bacteriostatic efficiency (>99.90%). Additionally, the particles produced by the combustion of TIBT-Cu are composed of active iodine substances and CuO particles, which can act synergistically against harmful microorganisms such as bacteria and viruses. This study provides a new perspective for the preparation of highly effective bactericidal materials.

Toxicity spectrum and risk factors for chemo-immunotherapy in locally advanced or metastatic lung cancer.

Chemo-immunotherapy has become the best first-line treatment for advanced lung cancer patients without oncogenic drivers. However, it may also lead to an increased incidence and severity of treatment-related adverse events. In this retrospective study, lung cancer patients administrated with either anti-PD-1 or anti-PD-L1 treatment plus chemotherapy were included. Data on demographic characteristics, disease characteristics, treatment strategies, laboratory results, and clinical outcomes were collected from the Electronic Medical Records System and evaluation scales. Chi-square, univariate, and multivariate logistic regression analyses were used to identify the risk factors for immune-related adverse events (irAEs). A total of 116 patients were included in the study, and the majority experienced treatment-related adverse events. Adverse events of any grade were reported in 114 (98.3%) patients, with 73 (62.9%) experiencing Grade 3 or higher events. The most frequent adverse events were anemia (67.2%), decreased appetite (62.9%), and alopecia (53.4%). Fifty-four (46.6%) patients were diagnosed with irAEs, with hypothyroidism (28.4%) being the most commonly reported. Multivariable analysis demonstrated a significant correlation between the number of treatment cycles, elevated baseline levels of thyroid stimulating hormone (TSH) and interleukin-6 (IL-6) with irAEs (OR = 1.222, P = 0.009, OR = 1.945, P = 0.016, OR = 1.176, P = 0.004), and IL-6 was identified as a strong predictor of severe irAEs (OR = 1.084, P = 0.014). Our study demonstrated the safety of chemo-immunotherapy in lung cancer patients without additional toxicity. The number of treatment cycles, higher baseline levels of TSH and IL-6 were identified as potential clinical biomarkers for irAEs.

Cooperative Catalysis Mechanism of Brønsted and Lewis Acids from Al(OTf)3 with Methanol for ß-Cellobiose-to-Fructose Conversion: An Experimental and Theoretical Study.

Al-containing catalysts, e.g., Al(OTf)3, show good catalytic performance toward the conversion of cellulose to fructose in methanol solution. Here, we report the catalytic isomerization and alcoholysis mechanisms for the conversion of cellobiose to fructose at the PBE0/6-311++G(d,p), aug-cc-pVTZ theoretical level, combining the relevant experimental verifications of electrospray ionization mass spectrometry (ESI-MS), high-performance liquid chromatography (HPLC), and the attenuated total reflection-infrared (ATR-IR) spectra. From the alcoholysis of Al(OTf)3 in methanol solution, the catalytically active species involves both the [CH3OH2]+ Brønsted acid and the [Al(CH3O)(OTf)(CH3OH)4]+ Lewis acid. There are two reaction pathways, i.e., one through glucose (glycosidic bond cleavage followed by isomerization, w-G) and another through cellobiulose (isomerization followed by glycosidic bond cleavage, w-L). The Lewis acid ([Al(CH3O)(OTf)(CH3OH)4]+) is responsible for the aldose-ketose tautomerization, while the Brønsted acid ([CH3OH2]+) is in charge of ring-opening, ring-closure, and glycosidic bond cleavage. For both w-G and w-L, the rate-determining steps are related to the intramolecular [1,2]-H shift between C1-C2 for the aldose-ketose tautomerization catalyzed by the [Al(CH3O)(OTf)(CH3OH)4]+ species. The Lewis acid ([Al(CH3O)(OTf)(CH3OH)4]+) exhibits higher catalytic activity toward the aldose-ketose tautomerization of glycosyl-chain-glucose to glycosyl-chain-fructose than that of chain-glucose to chain-fructose. Besides, the Brønsted acid ([CH3OH2]+) shows higher catalytic activity toward the glycosidic bond cleavage of cellobiulose than that of cellobiose. Kinetically, the w-L pathway is predominant, whereas the w-G pathway is minor. The theoretically proposed mechanism has been experimentally testified. These insights may advance on the novel design of the catalytic system toward the conversion of cellulose to fructose.

Characterization of Methyl- and Acetyl-Ni Intermediates in Acetyl CoA Synthase Formed during Anaerobic CO2 and CO Fixation.

The Wood-Ljungdahl Pathway is a unique biological mechanism of carbon dioxide and carbon monoxide fixation proposed to operate through nickel-based organometallic intermediates. The most unusual steps in this metabolic cycle involve a complex of two distinct nickel-iron-sulfur proteins: CO dehydrogenase and acetyl-CoA synthase (CODH/ACS). Here, we describe the nickel-methyl and nickel-acetyl intermediates in ACS completing the characterization of all its proposed organometallic intermediates. A single nickel site (Nip) within the A cluster of ACS undergoes major geometric and redox changes as it transits the planar Nip, tetrahedral Nip-CO and planar Nip-Me and Nip-Ac intermediates. We propose that the Nip intermediates equilibrate among different redox states, driven by an electrochemical-chemical (EC) coupling process, and that geometric changes in the A-cluster linked to large protein conformational changes control entry of CO and the methyl group.

Effect of Different Fine Aggregate Characteristics on Fracture Toughness and Microstructure of Sand Concrete.

The fracture toughness of sand concrete is affected by aggregate characteristics. In order to study the possibility of exploiting tailings sand, available in large quantities in sand concrete, and find an approach to improve the toughness of sand concrete by selecting appropriate fine aggregate. Three distinct fine aggregates have been used. After characterizing the fine aggregate used, the mechanical properties were tested to characterize the toughness of sand concrete, the box-counting fractal dimensions were calculated to analyze the roughness of fracture surfaces, and the microstructure was tested to observe the path and width of microcracks and hydration products in sand concrete. The results show that the mineral composition of fine aggregates is close, but their fineness modulus, fine aggregate angularity (FAA) and gradation vary considerably; FAA has a significant impact on the fracture toughness of sand concrete. The higher the FAA value, the more resistant it is to crack expansion; with the FAA values of from 32 s to 44 s, the microcrack width in sand concrete was reduced from 0.25 um to 0.14 um; The fracture toughness and microstructure of sand concrete are also related to the gradation of fine aggregates, the better gradation can improve the performance of the interfacial transition zone (ITZ). The hydration products in the ITZ are also different because more reasonable gradation of aggregates reduces the voids between the fine aggregates and the cement paste and restrains the full growth of crystals. These results demonstrate that sand concrete has promising applications in the field of construction engineering.

Spin polarization assisted facile C-H activation by an S = 1 iron(iv)-bisimido complex: a comprehensive spectroscopic and theoretical investigation.

High valent iron terminal imido species (Fe[double bond, length as m-dash]NR) have been shown to be key reactive intermediates in C-H functionalization. However, the detailed structure-reactivity relationship in Fe[double bond, length as m-dash]NR species derived from studies of structurally well-characterized high-valent Fe[double bond, length as m-dash]NR complexes are still scarce, and the impact of imido N-substituents (electron-donating vs. electron-withdrawing) on their electronic structures and reactivities has not been thoroughly explored. In this study, we report spectroscopic and computational studies on a rare S = 1 iron(iv)-bisimido complex featuring trifluoromethyl groups on the imido N-substituents, [(IPr)Fe(NC(CF3)2Ph)2] (2), and two closely related S = 0 congeners bearing alkyl and aryl substituents, [(IPr)Fe(NC(CMe3)2Ph)2] (3) and [(IPr)Fe(NDipp)2] (1), respectively. Compared with 1 and 3, 2 exhibits a decreased Fe[double bond, length as m-dash]NR bond covalency due to the electron-withdrawing and the steric effect of the N-substituents, which further leads to a pseudo doubly degenerate ground electronic structure and spin polarization induced ß spin density on the imido nitrogens. This unique electronic structure, which differs from those of the well-studied Fe(iv)-oxido complexes and many previously reported Fe(iv)-imido complexes, provides both kinetic and thermodynamic advantages for facile C-H activation, compared to the S = 0 counterparts.

Coordination of sorbitol to Ga(OTf)3 in the liquid phase: an experimental and theoretical study.

In a solution of sorbitol (SBT) and Ga(OTf)3 compounds, the coordination of sorbitol (SBT) to [Ga(OTf)n]3-n (n = 0-3) has been investigated, using both ESI-MS spectra and density functional theory (DFT) calculations at the M06/6-311++g(d,p), aug-cc-pvtz level using a polarized continuum model (PCM-SMD). In sorbitol solution, the most stable conformer of sorbitol includes three intramolecular H-bonds, i.e., O2H⋯O4, O4H⋯O6, and O5H⋯O3. Through ESI-MS spectra, in a tetrahydrofuran solution of both SBT and Ga(OTf)3 compounds, five main species are observed, i.e., [Ga(SBT)]3+, [Ga(OTf)]2+, [Ga(SBT)2]3+, [Ga(OTf)(SBT)]2+, and [Ga(OTf)(SBT)2]2+. Through DFT calculations, in a solution of sorbitol (SBT) and Ga(OTf)3 compounds, the Ga3+ cation tends to form five six-coordination complexes, i.e., [Ga(η2O,O-OTf)3], [Ga(η3O2-O4-SBT)2]3+, [(η2O,O-OTf)Ga(η4O2-O5-SBT)]2+, [(η1O-OTf)(η2O2,O4-SBT)Ga(η3O3-O5-SBT)]2+, and [(η1O-OTf)(η2O,O-OTf)Ga(η3O3-O5-SBT)]+, which are in good agreement with the experimental observation of the ESI-MS spectra. For both [Ga(OTf)n]3-n (n = 1-3) and [Ga(SBT)m]3+ (m = 1, 2) complexes, the negative charge transfer from ligands to the Ga3+-center plays an important role in their stability, because of the strong polarization of the Ga3+ cation. For [Ga(OTf)n(SBT)m]3-n (n = 1, 2; m = 1, 2) complexes, the negative charge transfer from ligands to the Ga3+-center plays an essential role in their stability, accompanied by an electrostatic interaction between the Ga3+-center and ligands and/or spatial inclusion of ligands toward the Ga3+-center.

Seeing the cis-Dihydroxylating Intermediate: A Mononuclear Nonheme Iron-Peroxo Complex in cis-Dihydroxylation Reactions Modeling Rieske Dioxygenases.

The nature of reactive intermediates and the mechanism of the cis-dihydroxylation of arenes and olefins by Rieske dioxygenases and synthetic nonheme iron catalysts have been the topic of intense research over the past several decades. In this study, we report that a spectroscopically well characterized mononuclear nonheme iron(III)-peroxo complex reacts with olefins and naphthalene derivatives, yielding iron(III) cycloadducts that are isolated and characterized structurally and spectroscopically. Kinetics and product analysis reveal that the nonheme iron(III)-peroxo complex is a nucleophile that reacts with olefins and naphthalenes to yield cis-diol products. The present study reports the first example of the cis-dihydroxylation of substrates by a nonheme iron(III)-peroxo complex that yields cis-diol products.

Systematic Comparison and Rational Design of Theophylline Riboswitches for Effective Gene Repression.

Riboswitches are promising regulatory tools in synthetic biology. To date, 25 theophylline riboswitches have been developed for regulation of gene expression in bacteria. However, no one has systematically evaluated their regulatory effects. To promote efficient selection and application of theophylline riboswitches, we examined 25 theophylline riboswitches in Escherichia coli MG1655 and found that they varied widely in terms of activation/repression ratios and expression levels in the absence of theophylline. Of the 20 riboswitches that activate gene expression, only one exhibited a high activation ratio (63.6-fold) and low expression level without theophylline. Furthermore, none of the five riboswitches that repress gene expression were more than 2.0-fold efficient. To obtain an effective repression system, we rationally designed a novel theophylline riboswitch to control a downstream gene or genes by premature transcription termination. This riboswitch allowed theophylline-dependent downregulation of the TurboRFP reporter in a dose- and time-dependent manner. Its performance profile exceeded those of previously described repressive theophylline riboswitches. We then introduced as the second part a RepA tag (protein degradation tag) coding sequence fused at the 5'-terminal end of the turborfp gene, which further reduced protein level, while not reducing the repressive effect of the riboswitch. By combining two tandem theophylline riboswitches with a RepA tag, we constructed a regulatory cassette that represses the expression of the gene(s) of interest at both the transcriptional and posttranslational levels. This regulatory cassette can be used to repress the expression of any gene of interest and represents a crucial step toward harnessing theophylline riboswitches and expanding the synthetic biology toolbox. IMPORTANCE A variety of gene expression regulation tools with significant regulatory effects are essential for the construction of complex gene circuits in synthetic biology. Riboswitches have received wide attention due to their unique biochemical, structural, and genetic properties. Here, we have not only systematically and precisely characterized the regulatory properties of previously developed theophylline riboswitches but also engineered a novel repressive theophylline riboswitch acting at the transcriptional level. By introducing coding sequences of a tandem riboswitch and a RepA protein degradation tag at the 5' end of the reporter gene, we successfully constructed a simple and effective regulatory cassette for gene regulation. Our work provides useful biological components for the construction of synthetic biology gene circuits.