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Nickel oxide (NiOx) is a promising hole transport layer (HTL) to fabricate efficient and large-scale inverted perovskite solar cells (PSCs) due to its low cost and superior chemical stability. However, inverted PSCs based on NiOx are still lagging behind that of other HTL because of the poor quality of buried interface contact. Herein, a bidentate ligand, 4,6-bis (diphenylphosphino) phenoxazine (2DPP), is used to regulate the NiOx surface and perovskite buried interface. The diphosphine Lewis base in the 2DPP molecule can coordinate both with NiOx and lead ions at NiOx/perovskite interface, leading to high-quality perovskite films with minimized defects. It is found that the inverted PSCs with 2DPP-modified buried interface exhibit double advantages of being both fast charge extraction and reduced nonradiative recombination, which is a combination of multiple factors including favorable energetic alignment, improved interface contact and strong binding between NiOx/2DPP and perovskite. The optimal PSC based on 2DPP modification yields a champion power conversion efficiency (PCE) of 21.9%. The unencapsulated PSC maintains above 75% of its initial PCE in the air with a relative humidity (RH) of 30-40% for 1000 h.
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BACKGROUND: The purpose of this study was to perform a biomechanical analysis to compare different medial column fixation patterns for valgus pilon fractures in a case-based model. METHODS: Based on the fracture mapping, 48 valgus pilon fracture models were produced and assigned into four groups with different medial column fixation patterns: no fixation (NF), K-wires (KW), intramedullary screws (IS), and locking compression plate (LCP). Each group contained wedge-in and wedge-out subgroups. After fixing each specimen on the machine, gradually increased axial compressive loads were applied with a load speed of one millimeter per minute. The maximum peak force was set at 1500 N. Load-displacement curves were generated and the axial stiffness was calculated. Five different loads of 200 N, 400 N, 600 N, 800 N, 1000 N were selected for analysis. The specimen failure was defined as resultant loading displacement over 3 mm. RESULTS: For the wedge-out models, Group-IS showed less displacement (p < 0.001), higher axial stiffness (p < 0.01), and higher load to failure (p < 0.001) than Group-NF. Group-KW showed comparable displacement under loads of 200 N, 400 N and 600 N with both Group-IS and Group-LCP. For the wedge-in models, no statistical differences in displacement, axial stiffness, or load to failure were observed among the four groups. Overall, wedge-out models exhibited less axial stiffness than wedge-in models (all p < 0.01). CONCLUSIONS: Functional reduction with stable fixation of the medial column is essential for the biomechanical stability of valgus pilon fractures and medial column fixation provides the enough biomechanical stability for this kind of fracture in the combination of anterolateral fixation. In detail, the K-wires can provide a provisional stability at an early stage. Intramedullary screws are strong enough to provide the medial column stability as a definitive fixation. In future, this technique can be recommended for medial column fixation as a complement for holistic stability in high-energy valgus pilon fractures.
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Placas Óseas , Fracturas de la Tibia , Humanos , Fenómenos Biomecánicos , Fracturas de la Tibia/cirugía , Fracturas de la Tibia/diagnóstico por imagen , Fracturas de la Tibia/fisiopatología , Tornillos Óseos , Fijación Interna de Fracturas/métodos , Fijación Interna de Fracturas/instrumentación , Fracturas de Tobillo/cirugía , Fracturas de Tobillo/diagnóstico por imagen , Fracturas de Tobillo/fisiopatología , Hilos Ortopédicos , Masculino , Soporte de Peso , Femenino , Adulto , Persona de Mediana EdadRESUMEN
The purpose of this work is to assess the robustness of treatment plans when spot delivery errors were predicted with a machine learning (ML) model for intensity modulated proton therapy (IMPT). Over 6000 machine log files from delivered IMPT treatment plans were included in this study. From these log files, over 4.1 × $ \times \ $ 106 delivered proton spots were used to train the ML model. The presented model was tested and used to predict the spot position as well as the monitor units (MU) per spot, based on the original planning parameters. Two patient plans (one accelerated partial breast irradiation [APBI] and one ependymoma) were recalculated with the predicted spot position/MUs by the ML model and then were re-analyzed for robustness. Plans with ML predicted spots were less robust than the original clinical plans. In the APBI plan, dosimetric changes to the left lung and heart were not clinically relevant. In the ependymoma plan, the hot spot in the brainstem decreased and the hot spot in the cervical cord increased. Despite these differences, after robustness analysis, both ML spot delivery error plans resulted in >95% of the CTV receiving >95% of the prescription dose. The presented workflow has the potential benefit of including realistic spots information for plan quality checks in IMPT. This work demonstrates that in the two example plans, the plans were still robust when accounting for spot delivery errors as predicted by the ML model.
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Carcinoma de Pulmón de Células no Pequeñas , Neoplasias Pulmonares , Terapia de Protones , Radioterapia de Intensidad Modulada , Humanos , Terapia de Protones/métodos , Planificación de la Radioterapia Asistida por Computador/métodos , Dosificación Radioterapéutica , Radioterapia de Intensidad Modulada/métodosRESUMEN
OBJECTIVE: To observe the changes in semen parameters after COVID-19 infection and clarify its impact on male fertility. METHODS: We collected semen samples from 82 male patients infected with COVID-19 in the past 2 months (the infection group) and 14 normal healthy men (the control group), obtained their semen parameters and compared them between the two groups before and after COVID-19 infection. RESULTS: There were no statistically significant differences in the baseline semen parameters between the infection and control groups (P > 0.05), nor in the semen volume within the infection group before and after infection (P > 0.05). Compared with the normal controls, the patients showed significantly decreased sperm concentration, total sperm count, percentage of progressively motile sperm, sperm motility and percentage of morphologically normal sperm after COVID-19 infection (P < 0.05), which were reduced even more significantly in those with than in those without fever during infection (P < 0.05). No statistically significant difference was observed in the semen quality of the patients with normal body temperature before and after COVID-19 infection (P > 0.05). Spearman correlation analysis showed no significant correlation between semen parameters and the severity of fever during infection (P > 0.05). CONCLUSION: COVID-19 infection decreases the semen quality of the patient, and fever during infection is a significant influencing factor. The severity of fever, however, is not related to the reduction of semen quality.
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COVID-19 , Semen , Masculino , Humanos , Análisis de Semen , Estudios Retrospectivos , Motilidad Espermática , Recuento de Espermatozoides , EspermatozoidesRESUMEN
Core-shell photoanodes have shown great potential for photoelectrochemical (PEC) water oxidation. However, the construction of a high-quality interface between the core and shell, as well as a highly catalytic surface, remains a challenge. Herein, guided by computation, we present a BiVO4 photoanode coated with ZnCoFe polyphthalocyanine using pyrazine as a coordination agent. The bidirectional axial coordination of pyrazine plays a dual role by facilitating intimate interfacial contact between BiVO4 and ZnCoFe polyphthalocyanine, as well as regulating the electron density and spin configuration of metal sites in ZnCoFe phthalocyanine, thereby promoting the potential-limiting step of *OOH desorption. The resulting photoanode displayed a high photocurrent density of 5.7±0.1â mA cm-2 at 1.23â VRHE . This study introduces a new approach for constructing core-shell photoanodes, and uncovers the key role of pyrazine axial coordination in modulating the catalytic activity of metal phthalocyanine.
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Integrins are transmembrane proteins that transmit bi-directional signals across the cell membrane through global structural rearrangement among three different conformational states: bent, extended- closed, and extended-open conformations. However, the ß8 integrin is distinctive and may adopt only one conformation, that is, extended-closed conformation, with high affinity for ligands under physiological conditions, and may not transmit bi-directional signals like other integrin members. It is unclear how different ß8 domains affect its unique conformation and signaling. We swapped different domains of integrin ß3 with those of ß8 and investigated how they affected integrin ligand binding, global conformation, and outside-in signaling. We found that the ß8 epidermal growth factor (EGF) domains increased integrin ligand binding ability and contributed to its extended conformation. By comparison, the ß8 transmembrane and cytoplasmic domains had little effect on ligand binding or global conformation. The ß8 EGF and transmembrane domains did not affect integrin-mediated cell adhesion, cell spreading, focal adhesion formation, or colocalization of integrin with other proteins, but the cytoplasmic domain had a defective effect on outside-in signaling. Our results showed that different domains of ß8 play various roles on its unique conformation, ligand binding, and signaling, which are considered atypical among integrin members.
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Factor de Crecimiento Epidérmico , Integrinas , Integrinas/metabolismo , Factor de Crecimiento Epidérmico/metabolismo , Ligandos , Unión Proteica , Dominios Proteicos , Integrina beta3/genética , Integrina beta3/metabolismoRESUMEN
Researches were reported that respiratory diseases can lead to male infertility; however, it is unclear whether there is a relationship between pulmonary fibrosis (PF) and male infertility. This study examined the influence of PF on sperm quality and its mechanisms. The key signalling pathway of male infertility caused by PF was predicted based on bioinformatics research. After modelling, we evaluated semen quality. Real-time quantitative polymerase chain reaction and Western blotting were used to measure the protein and mRNA expression levels of phosphatidylinositol 3-kinase (PI3K), phosphorylation-protein kinase B (p-Akt) and B-cell lymphoma 2 (Bcl2) in rat testicular cells. Compared with group A (48.77 ± 4.67; 59.77 ± 4.79), the sperm concentration and total sperm viability of group B (8.44 ± 1.71; 15.39 ± 3.48) showed a downward trend (p < 0.05). Western blotting showed that the protein expressions of PI3K, p-Akt and Bcl2 in the testes of group B (0.30 ± 0.06; 0.27 ± 0.05; 0.15 ± 0.03) was significantly lower than those of group A (0.71 ± 0.07; 0.72 ± 0.06; 0.50 ± 0.06) (p < 0.05). The hypoxic environment induced by PF can inhibit the expression of PI3K, p-Akt and Bcl2 protein and eventually cause dysfunctional spermatogenesis.
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Proteínas Proto-Oncogénicas c-akt , Fibrosis Pulmonar , Animales , Masculino , Fosfatidilinositol 3-Quinasa/metabolismo , Fosfatidilinositol 3-Quinasa/farmacología , Fosfatidilinositol 3-Quinasas/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Fibrosis Pulmonar/metabolismo , Ratas , Análisis de Semen , EspermatozoidesRESUMEN
Depression and chronic prostatitis (CP) are two common diseases that affect the human population worldwide. Clinically, it has been demonstrated that andrological patients often simultaneously suffer from depression and CP. Prior investigations have established that depression acts as an independent risk factor for CP. Herein, we explored the correlation between depression and CP using bioinformatics tools and through animal experiments. The potential targets and signalling pathways involved in depression and CP were predicted using bioinformatics tool, while depression in the rat model was established through chronic restraint stress. The expression of the related proteins and mRNA was assessed by Western blotting and real-time fluorescence quantitative reverse transcription-polymerase chain reaction (RT-qPCR). Relative to those in the control rats, the protein contents of PI3K, p-Akt, and p-mTOR were lower in the model rats (p < 0.05). Similarly, the transcript levels of PI3K, Akt, and mTOR was also relatively lower in the model rats (p < 0.05). And PI3K/Akt agonists reduced inflammation in rat prostate tissue, accompanied by significant increases in the transcript and protein expression levels of PI3K, Akt, and mTOR. Thus, we proposed that depression model rats may induce CP as a result of mediation by the negative regulation of the PI3K/Akt/mTOR signalling network.
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Fosfatidilinositol 3-Quinasas , Prostatitis , Animales , Depresión/etiología , Humanos , Masculino , Fosfatidilinositol 3-Quinasas/metabolismo , Prostatitis/complicaciones , Proteínas Proto-Oncogénicas c-akt/metabolismo , Ratas , Ratas Sprague-Dawley , Serina-Treonina Quinasas TOR/metabolismoRESUMEN
PURPOSE: To evaluate the impact of a digital whiteboard system integrated with data from the oncology information system (OIS) on the urgency of physics quality assurance (QA) tasks in the radiation oncology department. METHODS: Quality check list (QCL) items in the Mosaiq OIS corresponding to eight discrete, sequential steps in the treatment planning process were created. A whiteboard to graphically display active QCLs automatically and in real time was implemented in March 2020 using R shiny. QCL data with completion status were collected in two 12-month time periods before and after whiteboard implementation: January 2019-December 2019 and July 2020-June 2021. For all plans requiring patient-specific QA, we recorded when each plan was available for physics QA and which treatments started the following day. We further classified those plans into four categories (urgency levels 1-4 with 4 being the most urgent) depending on how much time was available to perform QA. We compared the proportion of these next-day QAs in each category between time periods accounting for plan type, day of the week, and time of year. RESULTS: Overall QA numbers were similar between time periods with 797 and 765 QAs total. The total proportion of next-day QA decreased by 27% and the proportions of urgency levels 1 and 4 both showed significant decreases after whiteboard implementation of 29.2% and 54.9%, respectively ( p < 0.05 $p<0.05$ ). All plan types had reduced proportions of next-day QAs, especially nonstereotactic body radiation therapy (non-SBRT) (30.3% decrease, p < 0.05 $p < 0.05$ ). Fridays and the months of October-December had the highest proportion of next-day QAs but showed significant reductions of 19.1% and 40.6% in the proportion of next-day QAs, respectively ( p < 0.05 $p<0.05$ ). CONCLUSIONS: The integrated whiteboard system significantly reduced the proportion of last-minute physics work, increasing patient safety. Advantages of the integrated whiteboard were low cost, low overhead with automatic interface to the OIS, and concurrent user support.
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Física , Planificación de la Radioterapia Asistida por Computador , Humanos , Garantía de la Calidad de Atención de Salud , Programas InformáticosRESUMEN
Integrins are heterodimeric transmembrane proteins that play important roles in various biological processes. Most integrins serve as adhesion molecules and transmit bidirectional signaling across the cell membrane through global conformational changes from the bent closed to the extended open conformation. However, integrin ß8 is distinctive in structure and function. Its cytoplasmic domain lacks the conserved protein-binding sequence, which is important in transmitting inside-out signals, suggesting that integrin ß8 may have a different activation mechanism or lack such signaling. In addition, the ligand-binding or activating metal ion Mn2+ does not induce a global conformational change in integrin ß8 . It may have only one conformation, that is, an extended, closed conformation, but with high affinity for ligands under physiological conditions, and is, therefore, considered an atypical integrin member. The extended structure and high ligand-binding affinity of integrin αv ß8 make it ideal for encountering and binding ligands expressed on an opposing cell or in the extracellular matrix. In this review, we summarize the progress in integrin ß8 research with a focus on its distinctive function and structure among integrin members.
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Membrana Celular/metabolismo , Integrinas/metabolismo , Transducción de Señal/fisiología , Sitios de Unión , Humanos , Ligandos , Unión Proteica/fisiología , Conformación Proteica , Dominios Proteicos/fisiologíaRESUMEN
Many integrins transmit signals through global conformational changes. However, it is unclear whether integrin αv ß8 adopts a similar mechanism during integrin activation and signaling on the cell surface. Here, we showed that disulfide-bonded mutants, which prevented integrin αv ß8 lower leg dissociation, bound ligands with similar level as the wild-type protein, suggesting that αv ß8 ligand binding did not require lower leg disassociation. We further showed that the N-glycosylation mutant at the interface between the ß I and hybrid domains did not affect ligand binding, suggesting that the αv ß8 open headpiece was not present on the cell surface. We proposed that αv ß8 integrin may adopt only one state, that is, the extended conformation with a closed headpiece. Our results showed that two lower legs retained heterodimeric interfaces, and this association might be important for stabilizing integrin in the extended conformation. Therefore, αv ß8 may not transmit bidirectional signals across the plasma membrane but instead may serve as an anchoring site with high affinity and high accessibility for extracellular ligands.
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Integrinas/química , Células HEK293 , Humanos , Integrinas/genética , Integrinas/metabolismo , Ligandos , Mutación , Dominios ProteicosRESUMEN
In recent years, two-dimensional (2D) lead-free double perovskites have been attracting much attention because of their unique performance in photovoltaic solar cells and photocatalysis. Nonetheless, how thickness affects the photoelectric properties of lead-free double perovskite remains unclear. In this work, by means of density functional theory (DFT) with a spin orbit coupling (SOC) effect, we have investigated the electronic and optical properties systemically, including band structures, carrier mobility, optical absorption spectra, exciton-binding energies, band edges alignment and molecule adsorption performance of Cs2AgBiBr6 with different thicknesses. The calculated results revealed the thickness-induced band gap and optical performance for Cs2AgBiBr6. It shows a low band gap and outstanding optical absorption of visible and ultraviolet light. When the thickness is reduced to a monolayer, Cs2AgBiBr6 moves from an indirect band gap to a direct band gap. Moreover, the carrier mobility of Cs2AgBiBr6 is excellent and the exciton-binding energy increases with the decreased thickness. Importantly, an analysis of molecule adsorption and band edge alignment indicates that Cs2AgBiBr6 is prone to H2O adsorption and H2 desorption theoretically, which is conducive to the photocatalytic water splitting for hydrogen generation and other photovatalytic reactions. Our work suggests that Cs2AgBiBr6 is a potential candidate as a solar cell or a photocatalyst, and we provide theoretical explorations into reducing the layers of lead-free double perovskite materials to 2D atomic thickness for a better photocatalytic application, which can serve as guidelines for the design of excellent photocatalysts.
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CONTEXT: Achyranthes bidentata Blume (Amaranthaceae) (ABR) and semen vaccariae (SV) are used commonly in the clinical treatment of erectile dysfunction in males with diabetes mellitus (DMED) to strengthen the kidney and promote blood circulation, and often achieve good curative effects. OBJECTIVE: Explore mechanistic details of ABR + SV treatment against DMED. MATERIALS AND METHODS: Prediction of key targets by network pharmacology. A rat model of DM was established by streptozotocin injection (55 mg/kg). Apomorphine (100 µg/kg) was injected into rats to screen the DMED model. Group C (n = 6) and group M (n = 6) were gavaged with deionized water; group T (n = 6) was given Achyranthis bidentatae radix-semen vaccariae granule suspension (2.5 g/kg). It lasted 8 weeks. Real-time reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and western blotting (WB) were used to measure the expression of tissue-related proteins and mRNA. RESULTS: The predicted key targets are albumin (ALB), caspase-3 (CASP3), vascular endothelial growth factor A (VEGFA), angiotensin-converting enzyme (ACE), and endothelial nitric oxide synthase (eNOS). Compared with the M group (0.52 ± 0.04; 0.50 ± 0.03; 0.49 ± 0.02; 0.23 ± 0.03), CASP3, VEGFA, and ACE protein expression reduced in the T group (0.39 ± 0.06; 0.34 ± 0.03; 0.39 ± 0.03), and eNOS protein expression increased (0.34 ± 0.03). CONCLUSION: ABR + SV can improve erectile function in DMED rats. This study provides a potential mechanism for the treatment of DMED with ABR + SV and can benefit from more patients.
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Achyranthes , Diabetes Mellitus Experimental/tratamiento farmacológico , Disfunción Eréctil/tratamiento farmacológico , Farmacología en Red/métodos , Extractos Vegetales/uso terapéutico , Vaccaria , Animales , Diabetes Mellitus Experimental/patología , Modelos Animales de Enfermedad , Disfunción Eréctil/patología , Masculino , Extractos Vegetales/aislamiento & purificación , Ratas , Ratas Sprague-Dawley , Resultado del TratamientoRESUMEN
CONTEXT: The leech and centipede granules have good curative effects on many diabetic vascular diseases, including diabetes-induced erectile dysfunction (DIED). OBJECTIVE: To explore the effect of leech and centipede on erectile function in rats with diabetes-induced erectile dysfunction and its possible mechanism. MATERIALS AND METHODS: Thirty male Sprague-Dawley DIED rats were randomly divided into the model group (Group M), low-dose group (Group DD), high-dose group (Group DG) and tadalafil group (Group T) (n = 6); diabetic rats were induced by streptozotocin. Apomorphine was used to induce diabetic erectile dysfunction. The 'leech-centipede' granules (0.15 and 0.6 g/kg) were intragastrically administered in the DD and DG groups for 8 weeks. Blood glucose, serum insulin, testosterone, cGMP levels and protein expression changes were measured in each group. RESULTS: After 8 weeks, the erectile function of rats in the DG group significantly improved (1.26 ± 0.73). Penis tissue cGMP levels were higher in the DG group (1.48 ± 0.11) than in the M group (0.58 ± 0.15). Protein and mRNA expression levels of NOS were significantly higher (0.77 ± 0.05; 0.61 ± 0.02) but those of PDE5 (0.43 ± 0.05; 0.61 ± 0.03) were lower in the DG group than in the M group (0.37 ± 0.06; 0.51 ± 0.01; 0.78 ± 0.06; 0.81 ± 0.04). CONCLUSION: The leech-centipede can improve erectile dysfunction in DIED rats by regulating the expression of cGMP, NOS, and PDE5-related molecules in the PDE5 pathway. This study provides a potential mechanism for the treatment of DIED with leech-centipede.
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Diabetes Mellitus Experimental/tratamiento farmacológico , Disfunción Eréctil/tratamiento farmacológico , Extractos de Tejidos/farmacología , Animales , Glucemia/efectos de los fármacos , GMP Cíclico/metabolismo , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 5/metabolismo , Diabetes Mellitus Experimental/complicaciones , Disfunción Eréctil/etiología , Masculino , Óxido Nítrico Sintasa/metabolismo , Erección Peniana/efectos de los fármacos , Ratas , Ratas Sprague-Dawley , Transducción de Señal/efectos de los fármacos , EstreptozocinaRESUMEN
Objective: To investigate the impact of asthma on erectile function in rats and the expressions of related proteins. METHODS: Male rats were injected intraperitoneally with ovalbumin solution to induce asthma followed by subcutaneous injection of apomorphine at 100 µg/kg into the neck, and then observed for reduced frequency or loss of penile erection. Based on the results of observation, a model of asthma-induced ED (AED) was made in 6 of the animals, and another 6 normal male rats were taken as controls. The histomorphology of the corpus cavernosum was observed by HE staining, and the mRNA and protein expressions of phosphodiesterase 5 (PDE5) and nitric oxide synthase 3 (eNOS) in the testis tissue were determined by RT-qPCR and Western blot. RESULTS: Compared with the normal controls, the rats in the AED model group showed disorderly distribution of sinusoids and decreased density of endothelial and smooth muscle cells in the corpus cavernosum. The mRNA and protein expressions of PDE5 were significantly higher (P < 0.05), while those of eNOS remarkably lower in the AED model than in the control group (P < 0.05). CONCLUSIONS: Asthma can induce ED and change the histomorphology of the corpus cavernosum in rats by affecting the expressions of PDE5 and eNOS proteins.
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The introduction of oxygen vacancies (Ov) has been regarded as an effective method to enhance the catalytic performance of photoanodes in oxygen evolution reaction (OER). However, their stability under highly oxidizing environment is questionable but was rarely studied. Herein, NiFe-metal-organic framework (NiFe-MOFs) was conformally coated on oxygen-vacancy-rich BiVO4 (Ov-BiVO4 ) as the protective layer and cocatalyst, forming a core-shell structure with caffeic acid as bridging agent. The as-synthesized Ov-BiVO4 @NiFe-MOFs exhibits enhanced stability and a remarkable photocurrent density of 5.3±0.15â mA cm-2 at 1.23â V (vs. RHE). The reduced coordination number of Ni(Fe)-O and elevated valence state of Ni(Fe) in NiFe-MOFs layer greatly bolster OER, and the shifting of oxygen evolution sites from Ov-BiVO4 to NiFe-MOFs promotes Ov stabilization. Ovs can be effectively preserved by the coating of a thin NiFe-MOFs layer, leading to a photoanode of enhanced photocurrent and stability.
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It has been proposed that integrins adopt a bent, closed conformation with low ligand binding capability at resting state and switch into an extended, open conformation upon activation or interacting with extracellular matrix (ECM) ligand. In this study, we addressed how integrin conformational change at the ß genu affects ligand binding and signaling. We discovered that swapping of the ß3 epidermal growth factor-like (EGF) domain 1 and 2 with that of ß8 greatly promoted ligand binding in ß3 ß8 chimeras. Sequence alignment indicated that ß8 integrin uniquely lacks the interface between the EGF1 and 2. Disrupting this interface of the ß3 integrin increased integrin ligand binding. Furthermore, the interface is critical for integrin affinity regulation but not downstream outside-in signaling.
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Cadenas beta de Integrinas/química , Cadenas beta de Integrinas/metabolismo , Integrina beta3/química , Integrina beta3/metabolismo , Dominios Proteicos/fisiología , Sitios de Unión , Adhesión Celular , Quimera , Complejos de Coordinación/química , Cristalografía , Cisteína/química , Factor de Crecimiento Epidérmico/química , Matriz Extracelular/metabolismo , Células HEK293 , Humanos , Ligandos , Microscopía Confocal , Unión Proteica , Conformación Proteica , Transducción de SeñalRESUMEN
It has been proposed that integrins adopt a low affinity conformation under physiological conditions. Integrin can either be activated through cytoplasm or by binding of cations such as Mn2+ to the head domain. The cytoplasmic activation pathway, that is, inside-out signaling has been regarded as the physiological pathway for integrin activation. Integrin ß8 is important for neuron vascular development. However, due to the highly divergent cytoplasmic domain, this integrin probably does not rely on inside-out signaling for affinity regulation. We therefore hypothesized that the ß8 integrin uniquely assumes a constitutively high affinity state under physiological conditions. We discovered that ß8 indeed exhibited high binding to soluble vitronectin in the presence of Ca2+ and the ligand binding could not be further enhanced by addition of Mn2+ . The lower ectodomain stalk of the integrin, which is comprised by the integrin epidermal growth factor-like (I-EGF) domains and ßTD domain, is critical for this high affinity conformation. In addition, we found that unlike other integrins, Mg2+ at low concentration inhibited ß8 ligand binding. Mutagenesis studies indicated that ß8 integrin possesses a unique cation binding site which might contribute to the ligand binding affinity. Our study showed that both the ß8 lower ectodomain stalk and the head domain play an important role in its high affinity state under physiological conditions. J. Cell. Biochem. 118: 2044-2052, 2017. © 2016 Wiley Periodicals, Inc.
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Calcio/química , Integrina alfa5/química , Cadenas beta de Integrinas/química , Proteínas Recombinantes de Fusión/química , Vitronectina/química , Secuencia de Aminoácidos , Sitios de Unión , Calcio/metabolismo , Cationes Bivalentes , Expresión Génica , Células HEK293 , Humanos , Integrina alfa5/genética , Integrina alfa5/metabolismo , Cadenas beta de Integrinas/genética , Cadenas beta de Integrinas/metabolismo , Ligandos , Magnesio/química , Magnesio/metabolismo , Manganeso/química , Manganeso/metabolismo , Mutación , Unión Proteica , Dominios Proteicos , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/metabolismo , Alineación de Secuencia , Homología de Secuencia de Aminoácido , Solubilidad , Soluciones , Vitronectina/genética , Vitronectina/metabolismoRESUMEN
Structures of intact receptors with single-pass transmembrane domains are essential to understand how extracellular and cytoplasmic domains regulate association and signaling through transmembrane domains. A chemical and computational method to determine structures of the membrane regions of such receptors on the cell surface is developed here and validated with glycophorin A. An integrin heterodimer structure reveals association over most of the lengths of the alpha and beta transmembrane domains and shows that the principles governing association of hetero and homo transmembrane dimers differ. A turn at the Gly of the juxtamembrane GFFKR motif caps the alpha TM helix and brings the two Phe of GFFKR into the alpha/beta interface. A juxtamembrane Lys residue in beta also has an important role in the interface. The structure shows how transmembrane association/dissociation regulates integrin signaling. A joint ectodomain and membrane structure shows that substantial flexibility between the extracellular and TM domains is compatible with TM signaling.
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Membrana Celular/química , Complejo GPIIb-IIIa de Glicoproteína Plaquetaria/química , Secuencia de Aminoácidos , Membrana Celular/metabolismo , Humanos , Modelos Moleculares , Datos de Secuencia Molecular , Mutación , Resonancia Magnética Nuclear Biomolecular , Complejo GPIIb-IIIa de Glicoproteína Plaquetaria/metabolismo , Estructura Terciaria de Proteína , Transducción de SeñalRESUMEN
Antifreeze proteins make up a class of ice-binding proteins (IBPs) that are possessed and expressed by certain cold-adapted organisms to enhance their freezing tolerance. Here we report the biophysical and functional characterization of an IBP discovered in a bacterium recovered from a deep glacial ice core drilled at Vostok Station, Antarctica (IBPv). Our study showed that the recombinant protein rIBPv exhibited a thermal hysteresis of 2 °C at concentrations of >50 µM, effectively inhibited ice recrystallization, and enhanced bacterial viability during freeze-thaw cycling. Circular dichroism scans indicated that rIBPv mainly consists of ß strands, and its denaturing temperature was 53.5 °C. Multiple-sequence alignment of homologous IBPs predicted that IBPv contains two ice-binding domains, a feature unique among known IBPs. To examine functional differences between the IBPv domains, each domain was cloned, expressed, and purified. The second domain (domain B) expressed greater ice binding activity. Data from thermal hysteresis and gel filtration assays supported the idea that the two domains cooperate to achieve a higher ice binding effect by forming heterodimers. However, physical linkage of the domains was not required for this effect.