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91.
The binding properties of the H5N1 influenza virus neuraminidase as inferred from molecular modeling
The avian influenza H5N1 virus has emerged as an important pathogen, causing severe disease in humans and posing a pandemic
threat. Substrate specificity is crucial for the virus to obtain the ability to spread from avian to human. Therefore, an
investigation of the binding properties of ligands at the molecular level is important for understanding the catalytic mechanism
of the avian influenza virus neuraminidase and for designing novel and specific inhibitors of H5N1 neuraminidase. Based on
the available crystal structure of H5N1, we have characterized the binding properties between sialic acid, methyl 3’sialyllactoside,
methyl 6’sialyllactoside and the H5N1 influenza virus neuraminidase using molecular docking and molecular dynamics simulations.
Obtained molecular dynamics trajectories were analyzed in terms of ligand conformations, N1-ligand interactions, and in terms
of loop flexibility. It was found that in the N1-SA complex the sialic acid ring undergoes a transition from the B
2,5 to the 2
C
5 conformation. However, in the N1-3SL and N1-6SL complexes sialic acid remained in the distorted boat conformation. The obtained
results indicate that 3SL has only weak interactions with the 150-loop, whereas the N1-6SL complex shows strong interactions.
Most of the differences arise from the various conformations around the glycosidic linkage, between the sialic acid and galactose,
which facilitate the above interactions of 6SL with the enzyme, and as a consequence the interactions between the 150- and
430- loops. This finding suggests that the altered flexibility of loops in and around the active site is one of the reasons
why the avian N1 preferentially cleaves sialic acid from α-(2-3)-Gal glycoconjugates over α-(2-6)-Gal. These molecular modeling
results are consistent with available experimental results on the specificity of N1. 相似文献
92.
Xu K Liu L Saad OM Baudys J Williams L Leipold D Shen B Raab H Junutula JR Kim A Kaur S 《Analytical biochemistry》2011,(1):56-66
Antibody–drug conjugates (ADCs) are designed to facilitate the targeted delivery of cytotoxic drugs to improve their tumor fighting effects and minimize systemic toxicity. However, efficacy and safety can potentially be compromised due to the release of conjugated drugs from the ADC with time while in circulation, resulting in changes in the drug-to-antibody ratio (DAR). Current understanding of this process is limited because existing methods such as immunoassays fail to distinguish ADCs with different DARs. Here we demonstrate a novel method with bead-based affinity capture and capillary liquid chromatography–mass spectrometry to allow direct measurement of drug release by quantifying DAR distributions of the ADC in plasma/serum. This method successfully identified individual intact conjugated antibody species produced due to drug loss from ADCs (e.g., an engineered site-specific anti-MUC16 THIOMAB–drug conjugate) and measured the corresponding DAR distributions in vitro and in vivo. Information obtained can provide insights into the mechanisms involved in drug loss and help to optimize ADC therapeutics. Other potential applications of the method may include characterization of posttranslational modifications, protein adducts, and immunogenicity. 相似文献
93.
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95.
Diazotrophic Diversity and Distribution in the Tropical and Subtropical Atlantic Ocean 总被引:1,自引:1,他引:1
To understand the structure of marine diazotrophic communities in the tropical and subtropical Atlantic Ocean, the molecular diversity of the nifH gene was studied by nested PCR amplification using degenerate primers, followed by cloning and sequencing. Sequences of nifH genes were amplified from environmental DNA samples collected during three cruises (November-December 2000, March 2002, and October-November 2002) covering an area between 0 to 28.3°N and 56.6 to 18.5°W. A total of 170 unique sequences were recovered from 18 stations and 23 depths. Samples from the November-December 2000 cruise contained both unicellular and filamentous cyanobacterial nifH phylotypes, as well as γ-proteobacterial and cluster III sequences, so far only reported in the Pacific Ocean. In contrast, samples from the March 2002 cruise contained only phylotypes related to the uncultured group A unicellular cyanobacteria. The October-November 2002 cruise contained both filamentous and unicellular cyanobacterial and γ-proteobacterial sequences. Several sequences were identical at the nucleotide level to previously described environmental sequences from the Pacific Ocean, including group A sequences. The data suggest a community shift from filamentous cyanobacteria in surface waters to unicellular cyanobacteria and/or heterotrophic bacteria in deeper waters. With one exception, filamentous cyanobacterial nifH sequences were present within temperatures ranging between 26.5 and 30°C and where nitrate was undetectable. In contrast, nonfilamentous nifH sequences were found throughout a broader temperature range, 15 to 30°C, more often in waters with temperature of <26°C, and were sometimes recovered from waters with detectable nitrate concentrations. 相似文献
96.
Plitas G Chaudhry UI Kingham TP Raab JR DeMatteo RP 《Journal of immunology (Baltimore, Md. : 1950)》2007,178(7):4411-4416
NK dendritic cells (NKDC) are recently described immunologic cells that possess both lytic and Ag-presenting function and produce prolific quantities of IFN-gamma. The role of NKDC in innate immunity to bacterial infection is unknown. Because IFN-gamma is important in the immune response to Listeria monocytogenes (LM), we hypothesized that NKDC play a critical role during LM infection in mice. We found that LM increased the frequency and activation state of NKDC in vivo. Using in vivo intracellular cytokine analysis, we demonstrated that NKDC are a major source of early IFN-gamma during infection with LM. Adoptive transfer of wild-type NKDC into IFN-gamma-deficient recipients that were subsequently infected with LM decreased bacterial burden in the liver and spleen and prolonged survival. In contrast, NK cells were depleted early during LM infection, produced less IFN-gamma, and conferred less protection upon adoptive transfer into IFN-gamma-deficient mice. In vitro, LM induction of IFN-gamma secretion by NKDC depended on TLR9, in addition to IL-18 and IL-12. Our study establishes NKDC as innate immune responders to bacterial infection by virtue of their ability to secrete IFN-gamma. 相似文献
97.
Shrinking biosensors down to microscale dimensions enables increases in sensitivity and the ability to analyze minute samples such as the contents of individual cells. The goal of the present study is to create mobile microscale biosensors by attaching molecular beacons to microtubules and using kinesin molecular motors to transport these functionalized microtubules across two-dimensional surfaces. Previous work has shown that microfluidic channels can be functionalized with kinesin motors such that microtubules can be transported and directed through these channels without the need for external power or pressure-driven pumping. In this work, we show that molecular beacons can be attached to microtubules such that both the fluorescence reporting capability of the beacon and the motility of the microtubules are retained. These molecular beacon-functionalized microtubules were able to bind ssDNA target sequences, transport them across surfaces, and report their presence by an increase in fluorescence that was detected by fluorescence microscopy. This work is an important step toward creating hybrid microdevices for sensitive virus detection or analyzing mRNA profiles of individual cells. 相似文献
98.
Kain R Exner M Brandes R Ziebermayr R Cunningham D Alderson CA Davidovits A Raab I Jahn R Ashour O Spitzauer S Sunder-Plassmann G Fukuda M Klemm P Rees AJ Kerjaschki D 《Nature medicine》2008,14(10):1088-1096
Pauci-immune focal necrotizing glomerulonephritis (FNGN) is a severe inflammatory disease associated with autoantibodies to neutrophil cytoplasmic antigens (ANCA). Here we characterize autoantibodies to lysosomal membrane protein-2 (LAMP-2) and show that they are a new ANCA subtype present in almost all individuals with FNGN. Consequently, its prevalence is nearly twice that of the classical ANCAs that recognize myeloperoxidase or proteinase-3. Furthermore, antibodies to LAMP-2 cause pauci-immune FNGN when injected into rats, and a monoclonal antibody to human LAMP-2 (H4B4) induces apoptosis of human microvascular endothelium in vitro. The autoantibodies in individuals with pauci-immune FNGN commonly recognize a human LAMP-2 epitope (designated P(41-49)) with 100% homology to the bacterial adhesin FimH, with which they cross-react. Rats immunized with FimH develop pauci-immune FNGN and also develop antibodies to rat and human LAMP-2. Finally, we show that infections with fimbriated pathogens are common before the onset of FNGN. Thus, FimH-triggered autoimmunity to LAMP-2 provides a previously undescribed clinically relevant molecular mechanism for the development of pauci-immune FNGN. 相似文献
99.
Junutula JR Raab H Clark S Bhakta S Leipold DD Weir S Chen Y Simpson M Tsai SP Dennis MS Lu Y Meng YG Ng C Yang J Lee CC Duenas E Gorrell J Katta V Kim A McDorman K Flagella K Venook R Ross S Spencer SD Lee Wong W Lowman HB Vandlen R Sliwkowski MX Scheller RH Polakis P Mallet W 《Nature biotechnology》2008,26(8):925-932
Antibody-drug conjugates enhance the antitumor effects of antibodies and reduce adverse systemic effects of potent cytotoxic drugs. However, conventional drug conjugation strategies yield heterogenous conjugates with relatively narrow therapeutic index (maximum tolerated dose/curative dose). Using leads from our previously described phage display-based method to predict suitable conjugation sites, we engineered cysteine substitutions at positions on light and heavy chains that provide reactive thiol groups and do not perturb immunoglobulin folding and assembly, or alter antigen binding. When conjugated to monomethyl auristatin E, an antibody against the ovarian cancer antigen MUC16 is as efficacious as a conventional conjugate in mouse xenograft models. Moreover, it is tolerated at higher doses in rats and cynomolgus monkeys than the same conjugate prepared by conventional approaches. The favorable in vivo properties of the near-homogenous composition of this conjugate suggest that our strategy offers a general approach to retaining the antitumor efficacy of antibody-drug conjugates, while minimizing their systemic toxicity. 相似文献
100.
J Steinbuch AC van Dijk FHBM Schreuder MTB Truijman J Hendrikse PJ Nederkoorn A van der Lugt E Hermeling APG Hoeks WH Mess 《Cardiovascular ultrasound》2017,15(1):9