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Background: Lung cancer remains the most fatal tumorous disease in the worldwide. Among that, lung adenocarcinoma (LUAD) was the most common histological type. A precise and concise prognostic model was urgently needed of LUAD. We developed a 23-gene signature for prognosis prediction based on EMT, immune and stromal datasets.Methods: Univariate Cox regression analysis was performed to select genes which were significantly associated with overall survival (OS) of the TCGA LUAD cohorts. LASSO regression and multivariate Cox regression analysis was used to build the multi-gene signature. Enrichment analyses and a protein-protein interactions (PPI) network were performed to show the interaction and functions of the signature. A nomogram was developed based on risk score and other clinical features. Predictive performance of the signature was externally validated in two independent datasets from Gene Expression Omnibus (GSE37745 and GSE13213).Results: A total of 1334 EMT, immune and stromal associated genes were obtained. After LASSO regression and multivariate Cox regression analysis, a 23-gene signature for risk stratification was built. K-M curves showed that the patients with high risk had a poorer outcome. Finally, a nomogram was built to predict prognosis. The predictive performance of the 23-gene signature was confirmed in internal and external validation.Conclusion: We developed and verified a 23-gene signature based on EMT, immune and stromal gene sets. It provided a convenient and concise tool for risk stratificationand individual medicine.

Background: Adenocarcinoma (AD) accounts for more than 30% of pulmonary carcinoma with the highest morbidity and mortality in the world. LncRNA regulates various biological processes and shows great potential to become a therapeutic target for AD patients. Methods: In this research, we combined normal lung tissue sample from the Genotype-Tissue Expression (GTEx) database and AD cohort in the Cancer Genome Atlas (TCGA) project to screen the significant Long non-coding RNAs (lncRNA). Besides, Gene Expression Omnibus (GEO) database and in vitro experiment were applied to subsequent verification. Findings: After screening, we selected PCPB1-AS1 as the core of follow-up research. Bioinformatic analyses indicated that PCBP1-AS1 specifically influenced intratumoral infiltration of T regulatory cell (Treg), which was verified by the experiments based on our cohort and AD cell lines. Meanwhile, ceRNA network suggested that RORA might be potential target of PCBP1-AS1 to regulate the Treg infiltration. In addition, the prediction of response to immune checkpoint inhibitors (ICI) treatment indicated PCBP1-AS1 expression level was able to become reference for ICIs treatment application. Interpretation: The compelling evidence demonstrated he rich biological functions of PCBP1-AS1 and its strong possibility to become the potential target for AD therapy. Funding Statement: National Natural Science Foundation (81672288), Focus on Research and Development Plan in Shandong Province (2018GSF118028) and Natural Science Foundation of Shandong Province (ZR2019PH002). Declaration of Interests: The authors declare that they have no conflict of interest. Ethics Approval Statement: This research was approved by Ethics Review Committee of Shandong Provincial Hospital, and was performed based on the Declaration of Helsinki and Good Clinical Practice guidelines, as defined by the International Conference on Harmonisation.

NIN/RPN12 binding protein 1 (NOB1p) encoded by NOB1 has been found to be an essential factor in 26S proteasome biogenesis which participates in protein degradation. However, the functions of NOB1 in non-small cell lung cancer cells are largely unknown. In the present study, lentivirus-mediated NOB1 shRNA transfection in two non-small cell lung cancer cell lines (A549 and H1299) was accomplished, as determined by fluorescence imaging. Downregulation of NOB1 expression was confirmed by real-time PCR and western blotting. NOB1 silencing resulted in a significant decline in the proliferation and colony formation capability of non-small cell lung cancer cells. Moreover, flow cytometry showed that A549 cells were arrested in the G0/G1 phase of the cell cycle after NOB1 suppression. Furthermore, depletion of NOB1 resulted in a significant decrease in CDK4 and cyclin D1 expression. These results suggest that NOB1 may act as an important regulator in non-small cell lung cancer growth and could be a therapeutic target of non‑small cell lung cancer.

We have determined by X-ray crystallography the structures of three dinuclear zirconium(IV) complexes containing the heptadentate ligand dhpta (where H(5)dhpta = 1,3-diamino-2-propanol-N,N,N',N'-tetraacetic acid, 1) and different countercations: K(2)[Zr(2)(dhpta)(2)].5H(2)O (2.5H(2)O), Na(2)[Zr(2)(dhpta)(2)].7H(2)O.C(2)H(5)OH (3.7H(2)O.C(2)H(5)OH), and Cs(2)[Zr(2)(dhpta)(2)].H(5)O(2).Cl.4H(2)O (4.H(5)O(2).Cl.4H(2)O). In the K(I) complex 2, crystallized from water, the two Zr(IV) ions are 3.5973(4) A apart and bridged via two alkoxo groups (average Zr-O 2.165 A). Each Zr(IV) is eight-coordinate and also bound to two N atoms (average Zr-N 2.448 A), and four carboxylate O atoms (average Zr-O 2.148 A). The two dhpta ligands in the dinuclear unit have different conformations. One face of the complex contains an array of 14 oxygen atoms and interacts strongly with the two K(I) ions, one of which is 6-coordinate, the other 8-coordinate, which are 3.922(4) A apart and bridged by a carboxylate O and by two water molecules. The structures of the dinuclear anion [Zr(2)(dhpta)(2)](2-) in the Na(I) complex 3 and in the Cs(I) complex 4 are essentially identical to that found in complex 2, although the alkali metal ions coordinate differently to the oxygen-rich face. All Zr(IV) ions have a distorted triangulated dodecahedral geometry. Although the crystal structure of complex 2 does not indicate the presence of acidic protons, in 4 an [H(5)O(2)](+) unit is strongly H-bonded to an oxygen atom of a coordinated carboxylate group. 1D and 2D (1)H and (13)C NMR spectroscopic and potentiometric studies reveal two deprotonations with pK(a) values of 9.0 and 10.0. At low pH, two carboxylate groups appear to undergo protonation accompanied by chelate ring-opening, and the complex exhibits dynamic fluxional behavior in which the two magnetically nonequivalent dhpta ligands exchange at a rate of 11 s(-1) at pH 3.30, 298 K, as determined from 2D EXSY NMR studies. Ligand interchange is not observed at high pH (>11). The same crystals of complex 2 were obtained from solutions at pH 3 or 12. The dynamic configurational change is therefore mediated by the aqueous solvent.

Human serum apotransferrin (hTF) binds to Zr(IV) slowly in the presence of nitrilotriacetate (NTA), citrate or ethylenediaminetetraacetate (EDTA) as donor ligands. For Zr(NTA)(2)(2-) as donor, equilibrium was reached in ca. 2 h (pH 7.4, 298 K, 10 mM Hepes, 5 mM bicarbonate) and full loading of the N- and C-lobe sites was achievable to give Zr(2)-hTF. (13)C NMR data suggest that carbonate can bind as a synergistic anion. (1)H and 2D [(1)H,(13)C] (using epsilon-[(13)C]Met-hTF) NMR studies show that there is little lobe-selectively in the order of Zr(IV) uptake. Fe(III) displaced Zr(IV) from the C-lobe of Zr(2)-hTF first, followed by the N-lobe. However, in the presence of a large excess of NTA, Zr(IV) binds to the N-lobe of holo-hTF (Fe(2)-hTF) first followed by the C-lobe. The (1)H and (13)C NMR chemical shift changes for epsilon-[(13)CH(3)] of Met464, which is close to the C-lobe site, are quite distinct from those observed previously for Al(III), Fe(III), Ti(IV), Ga(III) and Bi(III) binding to hTF, suggesting that Zr(IV) binding may not induce lobe closure [as observed previously for Hf(IV)]. This may affect receptor recognition and play a role in the different biological behaviour of Zr(IV) compared to Ti(IV).

The p K a values have been determined for eight of the nine histidine residues and the amino terminus of the N-lobe of human apo-transferrin (hTF/2N), and for seven of the nine histidine residues and the amino terminus of the protein Asp63Ser hTF/2N containing a mutation of the Fe 3+ -ligand Asp63 to Ser63. Calculations suggested that substitution of aspartate by serine would result in decreases of the p K a values of most of the histidine residues in the protein. This was found to be the case experimentally, and allowed assignment of the eCH resonance of His249. For the wild-type protein, the His residue with a p K a of 7.40 was assigned as His249, whereas for the mutant, no observable His residue had a p K a value higher than 6.9. The protonated form of His249 appears to be stabilised by interactions with Asp63, and the high p K a value may be critical for ensuring the release of iron at endosomal pH (5.5). The mutation lowered the apparent binding constant of hTF/2N for the synergistic anion oxalate from log K 4.0 to log K 3.3. 1 H NMR spectral changes induced by Ga 3+ binding to the mutant are compared to those observed for the wild-type protein.

The first observation of a redox process following a substitution reaction between a platinum(IV) complex K2PtCl6 with rabbit liver native Cd,Zn-MT is presented. The reaction features and products are studied by UV-visible and circular dichroism spectroscopy, chromatography, and X-ray photoelectron spectroscopic measurements. It is a significant complicated reaction comprising redox and substitution reactions. The reaction generates monomeric and dimeric products, and higher oligomers precipitate with intra- or intra- and intermolecular CyS-SCy linkages. Pt(IV) is reduced to Pt(II), which then binds to the monomeric and dimeric products, and may also bind to higher oligomers. The beta-cluster is more reactive than the alpha-cluster, and reacts first with K2PtCl6. Cd5Pt2 and Cd4Pt4 were found when native Cd,Zn-MT reacted with 2 and 4 molar equivalents of Pt(IV) for 2 h in which four Cd ions were located in the alpha-cluster. The amounts of Cd and Pt ions decreased in both monomeric and dimeric products when the reaction was prolonged and intramolecular CyS-SCy linkages increased. Besides the oligomers which precipitated, only dimeric products were formed when the reaction molar ratio of Pt(IV) to MT was more than 10:1. Cd3Pt6 and Cd1Pt8 were obtained when the reaction occurred for 2 and 72 h, respectively. The structure of the clusters may exist when native Cd, Zn-MT reacts with substoichiometric quantities of K2PtCl6 (0.5 K2PtCl6 per MT thiolate) for a short time (2 h), but may be partly disrupted with stoichiometric or excess quantities of K2PtCl6 (or = 0.5 K2PtCl6 per MT thiolate) for a long time. The disruption of the cluster structures results in an increase of the nonbridge thiolate and an increase of the binding sites to Pt ions. The mechanism of the antitumor activity and developing drug resistance of Pt(IV) complex drugs is discussed.

The ferric ion-binding protein A (FbpA), a member of transferrin superfamily, is a periplasmic iron transporter employed by many Gram-negative pathogens. Our experiments indicated copper(ii) could bind with Neisseria gonorrhoeae FbpA (NgFbpA), and the binding constant reached up to (8.7 ± 0.2) × 10(8) M(-1)via UV-vis titration. The crystal structure of recombinant Cu-NgFbpA at 2.1 Å revealed that the oxo-Cu2 clusters (dinuclear centres) assembled in the iron binding cleft and were bound to the two adjacent tyrosine residues (Y195 and Y196) of the protein, two Cu ions coordinated with two tyrosines, Y195 and Y196, respectively, which was different from the binding model of Fe ion with FbpA, in which Y195 and Y196 coordinated together with one Fe ion. While this was similar to the binding of Zr and Hf ion clusters, Y195 and Y196 coordinated with two metal ions and the μ-oxo-bridges linking the metal ions. Structural superimposition demonstrated that oxo-Cu2-NgFbpA still keeping an open conformation, similar to the apo-form of NgFbpA. The structure presented additional information towards an understanding of the function of FbpA, and provided a detailed binding model for FbpA protein with the possible metal ions in a biological system.

Transferrins transport Fe3+ and other metal ions in mononuclear-binding sites. We present the first evidence that a member of the transferrin superfamily is able to recognize multi-nuclear oxo-metal clusters, small mineral fragments that are the most abundant forms of many metals in the environment. We show that the ferric ion–binding protein from Neisseria gonorrhoeae (nFbp) readily binds clusters of Fe3+, Ti4+, Zr4+ or Hf4+ in solution. The 1.7 A resolution crystal structure of Hf–nFbp reveals three distinct types of clusters in an open, positively charged cleft between two hinged protein domains. A di-tyrosyl cluster nucleation motif (Tyr195-Tyr196) is situated at the bottom of this cleft and binds either a trinuclear oxo-Hf cluster, which is capped by phosphate, or a pentanuclear cluster, which in turn can be capped with phosphate. This first high-resolution structure of a protein–mineral interface suggests a novel metal-uptake mechanism and provides a model for protein-mediated mineralization/dissimilation, which plays a critical role in geochemical processes. NOTE: In the version of this article initially published online, the institution affiliations were assigned incorrectly because of a mistake that occurred during production. The correct affiliations for all authors are as follows: Dmitriy Alexeev1, Haizhong Zhu2, Maolin Guo2,3, Weiqing Zhong2,4, Dominic J.B. Hunter2, Weiping Yang2,3, Dominic J. Campopiano2 and Peter J. Sadler2. All of the footnotes (corrected) are as follows: 1Institute of Cell and Molecular Biology, Michael Swann Building, University of Edinburgh, Mayfield Road, Edinburgh EH9 3JR, UK; 2School of Chemistry, University of Edinburgh, West Mains Road, Edinburgh EH9 3JJ, UK; 3Current address: Department of Molecular Biology and Biochemistry, University of California, Irvine, California 92697, USA; and 4Current address: School of Pharmacy, Second Military Medicine University, Shanghai 200433, China. We apologize for any inconvenience this may have caused. This mistake has been corrected in the HTML and print version of the article.

This paper will be the first to discuss the in vivo and in vitro properties of a Pd(II) complex, K2PdCl4, interacting with metallothioneins (MTs). In vivo experiments revealed that intraperitoneal injections of K2PdCl4 into rabbits led to the simultaneous synthesis of Pd–MT in the kidney and Zn7MT in the liver. The renal Pd–MT complex contains 3.6 ± 0.3 Pd, 2.1 ± 0.2 Zn, and 1.0 ± 0.1 Cu per mole protein. It was found that pre-treatment with Zn(NO3)2 before K2PdCl4 injections significantly enhanced renal Pd–MT level. The same pre-treatment also increases hepatic Zn–MT levels. These results strongly suggest that Pd(II) ions can be bound in vivo by MT existing in the rabbit kidneys to form Pd–MT. Gel-filtration chromatographic studies after the incubation of either native Cd5Zn2MT2 or Zn7MT2 with K2PdCl4 in vitro demonstrate that Pd(II) ions promote the non-oxidative oligomerization of native MTs. Increasing the level of Pd(II) relative to MT led to a concomitant increase in the apparent yield of MT oligomers. At relatively low Pd–MT ratio, Pd(II) is found predominantly in the oligomers while the monomeric products are chiefly composed of the reactants, Cd5Zn2MT2 or Zn7MT2. Based on our experimental data, the mechanisms of the reactions between Pd(II) and MTs in vivo and in vitro are discussed.

The reactions of various rabbit MTs (Cd7Th, Zn7Th, Hg7Th, Bi7Th, Pb7Th) with EDTA were studied at different pH and EDTA concentrations, and were determined to be pseudo-first-order reactions. The reactions were all divided into two steps: one faster and EDTA-dependent, and the other EDTA-independent. The mechanism is also described in detail. Based on obtained results, the varied induction behaviors of MT formation for those metals are discussed.

We present a first‐principles study of elastic surface deformations, limits of atomic resolution, and of atomic‐scale friction in atomic force microscopy (AFM). In the case of a Pd AFM tip interacting with a graphite surface, we find that atomic resolution can be achieved in a narrow load range near ≊10−8 N (per Pd tip atom). For these loads, we determine the microscopic friction coefficient to be μ≊10−2.