PNNL-13501 - Pacific Northwest National Laboratory

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in the Environmental Molecular Sciences Laboratory. In addition to the resources at MSCF, the DOE supercomputers at the National Energy Research Scientific Computing Center, Lawrence Berkeley National Laboratory were also used to perform parts of the computations. Results and Accomplishments Molecular dynamics simulations for p21Ras alone and in complex with p120GAP were successfully completed during the last year. In the p21Ras complexed with p120GAP, only the Ras binding domain of GAP was included in the simulations because only that part of the protein is resolved in the x-ray experiments. The molecular dynamics simulation systems were carefully set up by first studying the protonation states (pKa) of the protein residues and of the bound GTP. Our pKa calculations showed that the protonation state of GTP bound to p21Ras alone and bound to p21Ras:p120GAP complex may be different. It was computed that the terminal γ-phosphate group of GTP would be protonated when p21Ras is alone, and it is unprotonated when p21Ras forms a complex with p120GAP. Since it would change the initial proton transfer to the GTP step of the reaction, this difference in the protonation states implies that the reaction mechanism of GTP to GDP hydrolysis may be different for the p21Ras alone and p21Ras:p120GAP systems. Gln61 is believed to play an important role in the structural stabilization of the catalytic site. In the molecular dynamics simulation of p21Ras alone, Gln61 stays fully solvent exposed as in the experimental x-ray structures. Gln61 is far from GTP, and therefore, most likely does not get involved in the hydrolysis reaction directly. It, however, contributes to the structural stability of the local structure around the GTP binding pocket. In contrast to the p21Ras alone system, Gln61 is close to the GTP in the p21Ras:p120GAP complex and interacts with GTP through a bridging water. It was observed in the molecular dynamics simulations that Gln61 might be occupying a position different than what was reported in the x-ray structure. In its observed configuration, it interacts with the GTP less specifically and contributes to the hydrolysis reaction by helping to align the bridging water in the proper orientation for the nucleophilic attack. The x-ray crystal structure of the p21Ras:p120GAP complex contains a water between Gln61 and GTP. This water (we named it nucleophilic water) is believed to be 142 FY 2000 Laboratory Directed Research and Development Annual Report the precursor of the nucleophilic hydroxyl that attacks the γ-phosphate of GTP during the hydrolysis. In the crystal structure, the nucleophilic water interacts with the sidechain carbonyl group of Gln61. During our simulation, there was a structural rearrangement of the GTP binding pocket of the protein complex. During the rearrangement, the nucleophilic water lost its interaction with the carbonyl group of Gln61 and started to interact with the NH2 sidechain group of the same residue. The structure after the rearrangement supports the earlier studies that Gln61 is probably not the general base in the hydrolysis reaction. The arginine residue (arginine finger) of p120GAP pointing toward GTP may increase the GTPase activity of Ras by stabilizing the transition state through favorable electrostatic interactions. This hypothesis successfully predicts that the GTPase activity of p21Ras increases when it binds p120GAP and that the activity of Gα (with its existing arginine finger) is higher than that of p21Ras. It, however, cannot explain why the GTPase activity of p21Ras:p120GAP is several orders of magnitude higher than that of Gα even though both members of the systems contain the arginine finger. In our p21Ras:p120GAP molecular dynamics simulations, we have observed a dominant structure of the GTP binding pocket. This dominant configuration is quite different from the x-ray crystal structure of the same complex. In this configuration, the nucleophilic water moves into a position such that its motion, orientation, and structural stability is controlled by its interactions with the backbone carbonyl of the arginine finger (Arg789 of p120GAP) and with the NH2 sidechain group of Gln61. In the corresponding structure of Gα, the water molecule does not interact with the backbone of the arginine finger residue. Based on our observations, we have postulated that, in addition to its transition state stabilization role as in the Gα system, the arginine finger of p120GAP also controls the orientation of the nucleophilic water while initiating the hydrolysis reaction. It is the single-handed operation of orienting the attacking water and the transition state stabilization by the same residue that makes the GTPase activity of p21Ras:p120GAP complex higher than that of the Gα system. Summary and Conclusions Protonation state and molecular simulation computations of the p21Ras and the p21Ras:p120GAP complex were successfully completed. We found that the protonation state of the bound GTP may be different in these two
systems. The dominant structure of the GTP binding site in the Ras:RasGAP complex was different from the corresponding x-ray structure. The dominant structure, which has not been reported before in the literature, is consistent with several key experimental results. These studies provide understanding the function of several key residues of the involved proteins. Next, we will determine the energetics and the pathways of the hydrolysis reaction. These studies will be performed by using quantum mechanical or hybrid quantum mechanical-classical mechanical approaches. Publication Resat H, TP Straatsma, DA Dixon, and JH Miller. “The arginine finger of RasGAP helps Gln61 align the nucleophilic water in GAP-stimulated hydrolysis of GTP.” Science (in preparation). Computational Science and Engineering 143
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Laboratory Directed Research and De
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Laboratory Directed Research and De
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Computational Science and Engineeri
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Policy and Management Sciences Asse
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Introduction Department of Energy O
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5. After reviews by the Technical a
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• The Technical Council peer revi
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methods applicable to combustion, s
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Summary Each national laboratory mu
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Study Control Number: PN0004/1411 A
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Table 1. Positive and negative ions
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m/z Arbitrary Units Figure 1. Mass
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introduce low-volatility compounds
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arrangement, but the charge is reta
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two liquid chromatographic gradient
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Integrated Microfabricated Devices
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Matrix Assisted Laser Desorption/Io
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Molecular Beam Synthesis and Charac
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temperature distribution of the des
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expected to result in significant a
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Study Control Number: PN99069/1397
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Seuss DT and KA Prather. 1999. “M
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Analysis of Receptor-Modulated Ion
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laboratory (Lu and Xie 1997; Lu et
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Automated DNA Fingerprinting Microa
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Comparison of 4 Xanthomonas Isolate
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Approach Hematopoietic (bone marrow
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Anti-Human lgG Human lgG Protein G
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Characterization of Global Gene Reg
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PCR products for immobilization on
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identify novel proteins of importan
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Immunoprecipitaton of tyrosinephosp
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Coupled NMR and Confocal Microscopy
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In the optical microscopy image, th
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Development and Applications of a M
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Figure 3. Hepa-1 cells undergoing a
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cellular processing (such as post-t
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8. Initial demonstration of an off-
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expression systems for several year
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accumulation of protein products, i
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Fungal Conversion of Agricultural W
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Fungal Conversion of Waste Glucose/
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Summary and Conclusions We demonstr
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are shown in Figure 1(a) and 1(b),
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Anchorage-Independent Growth (% con
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selection of seedlings demonstratin
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NMR-Based Structural Genomics Micha
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Earth System Science
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the lateral extent of the plume so
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Biogeochemical Processes and Microb
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Analyses of populations of viable a
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The first task was to establish fiv
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purged for 2 minutes. The gas sampl
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developing code architecture to min
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Berkowitz, CM. June 2000. “Atmosp
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environmental monitoring, 2) portab
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corresponded to a current density o
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Enhancing Emissions Pre-Processing
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Environmental Fate and Effects of D
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nitrogen and unpredictable eutrophi
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Figure 1. Ordinary kriging of 2 m d
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precipitation of calcite occurred i
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Interrelationships Between Processe
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phenanthrene resistant fraction con
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injection of an immiscible gas phas
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still some key kinetic issues that
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Application of a Crop Model The res
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The Nature, Occurrence, and Origin
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Particle number concentration, 1/cm
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geometry optimized AlOOH and FeOOH
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allowable parameters using thermody
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TCE + 3H + + 3Fe 2+ Fe)=> acetylene
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Use of Shear-Thinning Fluids for In
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concentration of 0.5% AMX was the m
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Roberts AL, LA Totten, WA Arnold, D
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tetra-scale computing, envisioned a
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Energy Technology and Management
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Figure 2. Setup for the second expe
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phase. The algorithms will be teste
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[Pb-212]/[Pb-212] final 100 10 1 0.
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Plasma Particle Dielectric Fiber El
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(species, sub-species, and sex), sp
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Figure 3. Relative risk of bone and
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Development of a Preliminary Physio
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Development of a Virtual Respirator
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Figure 3. Use of the chest cavity a
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Examination of the Use of Diazolumi
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Indoor Air Pathways to Nuclear, Che
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Source Building Release Observed re
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To illustrate the potential impact
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Non-Invasive Biological Monitoring
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Materials Science and Technology
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Figure 1. (a) - (c) Successive magn
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Component Fabrication and Assembly
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Development of a Low-Cost Photoelec
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elaxations, indicating the adequacy
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Promising chemical durability, as m
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Study Control Number : PN98028/1274
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High Power Density Solid Oxide Fuel
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Hybrid Plasma Process for Vacuum De
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Matson DW, PM Martin, WD Bennett, J
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We showed the proof-of-principle ap
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Bandgap Energy (eV) 2.9 2.85 2.8 2.
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Ultrathin Electrolyte Test Results
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Detailed Investigations on Hydrogen
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identified low energy step structur
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Development of Novel Photo-Catalyst
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-2 -1 Energy (eV) 0 1 2 Cu2O SrTiO3
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Here, we perform adsorption and des
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exceed those in the all-metal devic
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Intensity (a.u.) 0 100 200 300 400
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integrated voltage (V) 0.10 0.08 0.
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The first step was to implement thi
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Spontaneous Formation of Cu2O Nano-
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Actinide Chemistry at the Aqueous-M
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Figure 5. Close-up atomic force mic
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to 300°C and 400°C. Unfortunately
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Summary and Conclusions Transmutati
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Advanced Calibration/Registration T
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Figure 1a. Violet filter Figure 1b.
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Autonomous Ultrasonic Probe for Pro
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containing nitrous oxide, an optica
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Chemical Detection Using Cavity Enh
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Figure 3 is a color rendered simula
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appropriate study has been performe
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Exploitation of Conventional Chemic
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Figures 3 and 4 illustrate the impa
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enefits of using a modified spread-
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Single Channel Signal 0.0025 0.0020
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Therefore, in order to extract the
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A next-generation technology is nee
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Wind Dir. Figure 2. Negative ion co
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Volumetric Imaging of Materials by
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amplitude and peak frequency change
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Figure 3. 13 C NMR spectrum of corn
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Approach Two flow loops, one that o
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Modification of Ion Exchange Resins
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Permanganate Treatment for the Deco
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minimized by medium exchange. After
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Figure 1. Thermogravimetric analysi
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Validation and Scale-Up of Monosacc
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Concentration (g/100g solution) Con
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Analysis of High-Volume, Hyper-Dime
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ecent figure of merit values. Shoul
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Probabilistic Methods Development f
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a) Traditional PCA-based process co
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Acronyms and Abbreviations 2-D two-
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PNNL-13501 - Pacific Northwest National Laboratory

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