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Journal of Molecular Modeling Jun 2024The coherent electron/spin transport in azurin, a species of copper protein, was calculated based on the Landauer model. The research is motivated by the fast electron...
CONTEXT
The coherent electron/spin transport in azurin, a species of copper protein, was calculated based on the Landauer model. The research is motivated by the fast electron transport and spin selectivity/polarization in azurin, which have been reported in relation to the chiral-induced spin selectivity of the peptide structure. The calculated spin polarization of copper proteins was large. This phenomenon was strongly influenced by the spin density of the atoms in the ligand group, whereas the contribution of copper was negligible. The results suggest that spin polarization in copper proteins is enhanced by that of the ligand groups. The predicted spin polarization aligns primarily with the scanning tunneling microscope-based break-junction technique to study the electronic properties of single-molecule junctions.
METHODS
Computational techniques employed in this study are nonequilibrium Green's functions (NEGF) and density functional theory (DFT) based on the Landauer model, implemented using the QuantumATK software (Synopsys Inc.). The Perdew-Burke-Ernzerhof (PBE) exchange-correlation functional was adopted for spin-polarized generalized gradient approximation (SGGA). The valence atomic orbitals were constructed using the wavefunctions of the SIESTA package, which was based on the norm-conserving Troullier-Martins relativistic pseudopotentials for describing core electrons. The mesh used for real-space integration was 150 Ha.
Topics: Azurin; Copper; Models, Molecular; Electron Transport; Density Functional Theory
PubMed: 38890154
DOI: 10.1007/s00894-024-06025-9 -
Chemical Science May 2024Native chemical ligation (NCL) has been playing an increasingly important role in chemical protein synthesis (CPS). More efficient ligation methods that circumvent the...
Native chemical ligation (NCL) has been playing an increasingly important role in chemical protein synthesis (CPS). More efficient ligation methods that circumvent the requirement of a peptidyl thioester and thiol additive-which allow the following desulfurization or refolding in one pot-are urgently needed for the synthesis of more complex protein targets and in large quantities. Herein, we discover that the weak acyl donor peptidyl -acyl pyrazole can be activated by azole reagents like 3-methylpyrazole or imidazole to facilitate its ligation directly with an N-terminal cysteine peptide. As it requires no thioester or thiol additive, this ligation strategy can be conveniently combined with metal-free desulfurization (MFD) or oxidative protein folding to allow various one-pot protocols. The utility and generality of the strategy are showcased by the total synthesis of ubiquitin an N-to-C sequential ligation-MFD strategy, the semi-synthesis of the copper protein azurin, and the efficient assembly of a sulfated hirudin variant and the cyclotide kalata B1, all in a one-pot fashion.
PubMed: 38817582
DOI: 10.1039/d3sc06697e -
Clinical Laboratory May 2024The goal was to improve the clinical cognition of Ph-positive mixed phenotype acute leukemia and avoid misdiagnosis or delayed diagnosis.
BACKGROUND
The goal was to improve the clinical cognition of Ph-positive mixed phenotype acute leukemia and avoid misdiagnosis or delayed diagnosis.
METHODS
The clinical manifestations and laboratory results (bone marrow cell morphology, multiparameter flow cytometry, and cytogenetics) of a case of Ph-positive mixed phenotype acute leukemia were analyzed, and related literature was reviewed.
RESULTS
Blood routine: WBC 386.35 x 109/L, HGB 117.00 g/L, PLT 31 x 109/L; 80% of the original cells can be seen by artificial classification. Morphological examination of bone marrow cells showed that the proliferation of nucleated cells was obviously active, and the original cells accounted for 76%. The size of the original cells was somewhat uniform, most of the cells had less mass, were stained light grayish blue, the cytoplasm particles were not obvious, the nuclei were mostly round or quasi-round, some of them showed distortion and nuclear notch, and the chromatin was coarse. Some of the cells were rich in mass, small azurin granules were seen, the nuclei were regular, most of them were round, the chromatin was fine, the myeloperoxidase and esterase staining were negative, the eosinophils accounted for 2.5%, and the basophils accounted for 0.5%. Flow cytometry immunotyping: Two groups of abnormal cells were seen in the bone marrow. 1. A group included 12.32% of nuclear cells and showed abnormal myeloid primitive cell phenotype. Main expression: CD117, CD34, CD38, HLA-DR, CD33, CD64, CD123, weak expression: CD13, CD19. 2. The other group included 45.61% of the nuclear cells and had a B-lymphoblastic phenotype. Main expression: CD34, CD38, HLA-DR, CD123, CD19, CD10, CD9, cCD79a, TDT, weak expression of CD13, CD22. Mixed phenotype acute leukemia (M/B) immunophenotype was considered. Chromosome: 46,XY,t(9; 22)(q34;q11.2) [20]. BCR-ABL (P210) fusion gene was positive.
CONCLUSIONS
Mixed phenotype acute leukemia (MPAL) is a rare type of malignant hematologic disease. Its diagnosis is based on the comprehensive evaluation of bone marrow cell morphology, immunophenotype, molecular and cytogenetic features.
Topics: Humans; Phenotype; Flow Cytometry; Male; Immunophenotyping; Bone Marrow Cells; Philadelphia Chromosome; Leukemia, Biphenotypic, Acute; Leukemia; Adult; Female; Middle Aged
PubMed: 38747916
DOI: 10.7754/Clin.Lab.2023.231220 -
Analytical Methods : Advancing Methods... May 2024This work describes an analytical procedure, single particle-inductively coupled plasma-time-of-flight-mass spectrometry (SP-ICP-TOF-MS), that was developed to determine...
This work describes an analytical procedure, single particle-inductively coupled plasma-time-of-flight-mass spectrometry (SP-ICP-TOF-MS), that was developed to determine the platinum binding efficiency of protein-coated magnetic microparticles. SP-ICP-TOF-MS is advantageous due to its ability to quasi-simultaneously detect all nuclides (Li-Pu), allowing for both platinum and iron (composition of magnetic microparticles) to be measured concurrently. This method subsequently allows for the differentiation between bound and unbound platinum. The 1 μm magnetic microparticles were fully characterized for their iron concentration, particle concentration, and trace element composition by bulk digestion-ICP-MS and SP-ICP-TOF-MS. The results of both approaches agreed with the certificate values. Using the single particle methodology the platinum loading was quantified to be to 0.18 ± 0.02 fg per particle and 0.32 ± 0.02 fg per particle, for the streptavidin-coated and azurin-coated microparticles, respectively. Both streptavidin-coated and the azurin-coated microparticles had a particle-platinum association of >65%. Platinum bound samples were also analyzed bulk digestion-based ICP-MS. The bulk ICP-MS results overestimated platinum loading due to free platinum in the samples. This highlights the importance of single particle analysis for a closer inspection of platinum binding performance. The SP-ICP-TOF-MS approach offers advantages over typical bulk digestion methods by eliminating laborious sample preparation, enabling differentiation between bound/unbound platinum in a solution, and quantification of platinum on a particle-by-particle basis. The procedure presented here enables quantification of metal content per particle, which could be broadly implemented for other single particle applications.
Topics: Platinum; Mass Spectrometry; Microspheres; Iron; Streptavidin; Particle Size; Magnetite Nanoparticles
PubMed: 38639200
DOI: 10.1039/d4ay00268g -
Cell Reports. Medicine May 2024Bacteria-based therapies are powerful strategies for cancer therapy, yet their clinical application is limited by a lack of tunable genetic switches to safely regulate...
Bacteria-based therapies are powerful strategies for cancer therapy, yet their clinical application is limited by a lack of tunable genetic switches to safely regulate the local expression and release of therapeutic cargoes. Rapid advances in remote-control technologies have enabled precise control of biological processes in time and space. We developed therapeutically active engineered bacteria mediated by a sono-activatable integrated gene circuit based on the thermosensitive transcriptional repressor TlpA. Through promoter engineering and ribosome binding site screening, we achieved ultrasound (US)-induced protein expression and secretion in engineered bacteria with minimal noise and high induction efficiency. Specifically, delivered either intratumorally or intravenously, engineered bacteria colonizing tumors suppressed tumor growth through US-irradiation-induced release of the apoptotic protein azurin and an immune checkpoint inhibitor, a nanobody targeting programmed death-ligand 1, in different tumor mouse models. Beyond developing safe and high-performance designer bacteria for tumor therapy, our study illustrates a sonogenetics-controlled therapeutic platform that can be harnessed for bacteria-based precision medicine.
Topics: Animals; Mice; Humans; Neoplasms; Disease Models, Animal; Cell Line, Tumor; Female; B7-H1 Antigen; Immune Checkpoint Inhibitors; Escherichia coli
PubMed: 38608697
DOI: 10.1016/j.xcrm.2024.101513 -
Journal of Inorganic Biochemistry Jul 2024Motivated by the ambition to establish an enzyme-driven bioleaching pathway for copper extraction, properties of the Type-1 copper protein rusticyanin from...
Kinetic, electrochemical and spectral characterization of bacterial and archaeal rusticyanins; unexpected stability issues and consequences for applications in biotechnology.
Motivated by the ambition to establish an enzyme-driven bioleaching pathway for copper extraction, properties of the Type-1 copper protein rusticyanin from Acidithiobacillus ferrooxidans (AfR) were compared with those from an ancestral form of this enzyme (N0) and an archaeal enzyme identified in Ferroplasma acidiphilum (FaR). While both N0 and FaR show redox potentials similar to that of AfR their electron transport rates were significantly slower. The lack of a correlation between the redox potentials and electron transfer rates indicates that AfR and its associated electron transfer chain evolved to specifically facilitate the efficient conversion of the energy of iron oxidation to ATP formation. In F. acidiphilum this pathway is not as efficient unless it is up-regulated by an as of yet unknown mechanism. In addition, while the electrochemical properties of AfR were consistent with previous data, previously unreported behavior was found leading to a form that is associated with a partially unfolded form of the protein. The cyclic voltammetry (CV) response of AfR immobilized onto an electrode showed limited stability, which may be connected to the presence of the partially unfolded state of this protein. Insights gained in this study may thus inform the engineering of optimized rusticyanin variants for bioleaching processes as well as enzyme-catalyzed solubilization of copper-containing ores such as chalcopyrite.
Topics: Kinetics; Electrochemistry; Azurin; Actinobacteria; Thermoplasmales; Electron Spin Resonance Spectroscopy; Models, Molecular; Protein Structure, Tertiary; Iron; Oxidation-Reduction; Biotechnology; Protein Stability; Conserved Sequence
PubMed: 38593609
DOI: 10.1016/j.jinorgbio.2024.112539 -
The Journal of Physical Chemistry. B Apr 2024Secondary coordination sphere (SCS) interactions have been shown to play important roles in tuning reduction potentials and electron transfer (ET) properties of the Type...
Secondary coordination sphere (SCS) interactions have been shown to play important roles in tuning reduction potentials and electron transfer (ET) properties of the Type 1 copper proteins, but the precise roles of these interactions are not fully understood. In this work, we examined the influence of F114P, F114N, and N47S mutations in the SCS on the electronic structure of the T1 copper center in azurin (Az) by studying the hyperfine couplings of (i) histidine remote N nitrogens and (ii) the amide N using the two-dimensional (2D) pulsed electron paramagnetic resonance (EPR) technique HYSCORE (hyperfine sublevel correlation) combined with quantum mechanics/molecular mechanics (QM/MM) and DLPNO-CCSD calculations. Our data show that some components of hyperfine tensor and isotropic coupling in N47SAz and F114PAz (but not F114NAz) deviate by up to ∼±20% from WTAz, indicating that these mutations significantly influence the spin density distribution between the Cu site and coordinating ligands. Furthermore, our calculations support the assignment of N to the backbone amide of residue 47 (both in Asn and Ser variants). Since the spin density distributions play an important role in tuning the covalency of the Cu-Scys bond of Type 1 copper center that has been shown to be crucial in controlling the reduction potentials, this study provides additional insights into the electron spin factor in tuning the reduction potentials and ET properties.
Topics: Azurin; Copper; Nitrogen; Mutation; Electron Spin Resonance Spectroscopy; Amides; Alaska Natives
PubMed: 38564809
DOI: 10.1021/acs.jpcb.3c08194 -
Marine Drugs Jan 2024Biofilm is accountable for nosocomial infections and chronic illness, making it a serious economic and public health problem. , thanks to its ability to form biofilm and...
Biofilm is accountable for nosocomial infections and chronic illness, making it a serious economic and public health problem. , thanks to its ability to form biofilm and colonize biomaterials, represents the most frequent causative agent involved in biofilm-associated infections of medical devices. Therefore, the research of new molecules able to interfere with biofilm formation has a remarkable interest. In the present work, the attention was focused on sp. TAE6080, an Antarctic marine bacterium able to produce and secrete an effective antibiofilm compound. The molecule responsible for this activity was purified by an activity-guided approach and identified by LC-MS/MS. Results indicated the active protein was a periplasmic protein similar to the PAO1 azurin, named cold-azurin. The cold-azurin was recombinantly produced in and purified. The recombinant protein was able to impair attachment to the polystyrene surface and effectively prevent biofilm formation.
Topics: Pseudomonas; Azurin; Anti-Bacterial Agents; Antarctic Regions; Escherichia coli; Chromatography, Liquid; Tandem Mass Spectrometry; Biofilms; Pseudomonas aeruginosa; Staphylococcus epidermidis
PubMed: 38393032
DOI: 10.3390/md22020061 -
Journal of Inorganic Biochemistry May 2024Anthropogenic activities in agriculture and health use the antimicrobial properties of copper. This has led to copper accumulation in the environment and contributed to...
Anthropogenic activities in agriculture and health use the antimicrobial properties of copper. This has led to copper accumulation in the environment and contributed to the emergence of copper resistant microorganisms. Understanding bacterial copper homeostasis diversity is therefore highly relevant since it could provide valuable targets for novel antimicrobial treatments. The periplasmic CopI protein is a monodomain cupredoxin comprising several copper binding sites and is directly involved in copper resistance in bacteria. However, its structure and mechanism of action are yet to be determined. To study the different binding sites for cupric and cuprous ions and to understand their possible interactions, we have used mutants of the putative copper binding modules of CopI and spectroscopic methods to characterize their properties. We show that CopI is able to bind a cuprous ion in its central histidine/methionine-rich region and oxidize it thanks to its cupredoxin center. The resulting cupric ion can bind to a third site at the N-terminus of the protein. Nuclear magnetic resonance spectroscopy revealed that the central histidine/methionine-rich region exhibits a dynamic behavior and interacts with the cupredoxin binding region. CopI is therefore likely to participate in copper resistance by detoxifying the cuprous ions from the periplasm.
Topics: Copper; Histidine; Binding Sites; Methionine; Anti-Infective Agents; Ions; Azurin
PubMed: 38364337
DOI: 10.1016/j.jinorgbio.2024.112503 -
Molecular Pharmaceutics Feb 2024
PubMed: 38270493
DOI: 10.1021/acs.molpharmaceut.4c00011