dataesr/scientific-paragraphs-categorization huggingface dataset:
Receptor sublocalization was determined using rabbit monoclonal anti-GlyR ␣1 (1:400; Millipore) primary antibody and with goat anti-rabbit secondary antibody, conjugated with Alexa Fluor 488 (1:200; Invitrogen). Cell surface immunostaining was conducted using the same reagents, antibodies, and dilutions but carried out prior to paraformaldehyde fixation. Fixed cells were then quenched with 50 mM NH 4 Cl before mounting on glass slides. Cell images were acquired using a Zeiss LSM 710 confocal microscope with ZEN software. The master gain was kept constant to compare the expression of ␣1 GlyR mutants relative to wild type. Transfection was repeated three times. Molecular Modeling-Wild type and mutated forms of the human ␣1 GlyR were modeled by 50% homology (69% sequence coverage) with Protein Data Bank structure 3RHW, the glutamate-gated chloride channel receptor (␣ GluClR) from Caenorhabditis elegans  (22).
Using our multitemplate homology modeling pipeline, this was one of three Protein Data Bank homologues that were identified and used in the assembly of the ␣1 GlyR models, 3RHW (chain E), 1VRY (chain A), and 1MOT (chain A). The homology modeling pipeline was built with the Biskit structural bioinformatics platform  (23). Our pipeline workflow incorporates the NCBI tools platform  (24) , including the BLAST program for similarity searching of sequence databases. T-COFFEE  (25)  was used for alignment of the test sequence with the template, followed by iterations of the MODELLER-9. 11 program  (26)  to generate the final model structure. The Chimera program  (27)  was used for the viewing of models and generation of images.
RESULTS Mutation Analysis-A total of 68 probands with hyperekplexia were assessed for genetic variation in GLRA1 coding regions. All sequence variations were cross-referenced with the dbSNP database and our previous GLRA1 data sets (for recurrent mutations) and were regarded as probable mutations following exclusion from a panel of 100 control samples and the exome variants server. The screening revealed 19 mutations in 21 hyperekplexia probands (Table  1 ), a rate that is consistent with previous studies  (10, 28). Nine mutations were novel in the public domain, and three (one novel and two recurrent) mutations were present in more than one individual. Note that p. R414H has since been reported as a very rare variant in dbSNP (rs200130685), with a heterozygosity of 0. 002 and a minor allele frequency of 0.
Control recordings from MTSR-labeled p. R271C GlyRs are also shown. Again, these results are consistent with the fluorescence data presented in Fig. 1. Cell Surface Localization-Confocal microscopy of transfected cells immunostained with an ␣1 GlyR antibody was performed to analyze subcellular localization of wild type and mutated receptors. Permeabilized cells were used to determine the intracellular sublocalization of receptors with all constructs showing comparable intracellular expression (Fig. 6 , left panel of each image pair). Cell surface expression was visualized in nonpermeabilized cells. Cells transfected with wild type and the dominant mutations p. V280M and p. R414H showed immunoreactivity around the cellular circumference indicating localization at the surface (Fig. 6 , right panels), suggesting that these variants do not affect integration of GlyRs into the cell membrane.
To our knowledge, this is the first suggestion that a pLGIC receptor subunit may be functionally expressed without a TM4 domain. The human GlyR ␣4 subunit (Uniprot accession number Q5JXX5) has long been regarded as a pseudo-gene because it incorporates a stop codon that truncates the receptor prior to the TM4 domain  (3). The truncation occurs at residue Asp-383, which corresponds to Gln-382 in the ␣1 subunit. Because this is C-terminal of E375, it is possible that the ␣4 subunit may be incorporated into functional GlyRs, suggesting that it could exert a physiological role in modulating the functional properties of GlyRs. There may also be a case to reassess the pathophysiology of the other pLGIC late protein truncating events in epilepsy and related disorders, for example GABA A receptor mutations  (46).
S1 ) and describing the backbone motions  [71, 72]. Only the FEPs which were similar in the two trajectories of the protein in the same nucleotide-binding state were kept for the analysis of the allosteric communication, we named these FEPs the DATA. No information can be exploited from the other FEPs: they may or not be relevant to the conformational fluctuations of the protein (NO DATA set). With this approach, we were able to quantify how each FEP varies along the amino-acid sequence depending on the nucleotide-binding state of the protein and to relate the FEP modifications to biochemical data of wild-type or mutant E. coli DnaK chaperones. Indeed, the comparison between the DATA sets of FEPs for E. coli DnaK in different nucleotidebinding states lead finally to a subset of only 27 key dihedral angles involved in the conformational nucleotide-dependent transitions.
Hsp70 s occur in two main conformations to perform their chaperone function, we named closed conformation (with ADP bound to the NBD or without nucleotide = APO, Fig. 1 A ) and open conformation (with ATP bound to the NBD, Fig. 1 B ). In the nucleotide-free Hsp70 and in ADP-bound Hsp70, the SBD is assumed to be closed with low binding and release rate of the protein substrate. However, recent experiments shown that the complete closure of the SBD is not strictly required in the chaperone protein interactions  [81]. In absence of ATP, the SBD and the NBD are undocked and the inter-domain linker is exposed to solvent, as shown by low-resolution Small Angle X-ray Scattering (SAXS) data  [82, 83] , Fo ¨rster resonance energy transfer (FRET) data  [84, 85]  and by the two-domain Hsp70 NMR derived-structure (PDB ID: 2KHO)  [70]  (Fig.
1 A ). In ATP-bound Hsp70, the SBD is open with fast binding and release of the protein substrate, and the SBD and NBD are docked, as shown by SAXS data  [82, 83]  and FRET data  [84, 85]  and suggested by the X-ray structure of an ATP-Hsp110 homolog  [86, 87]  and by a new open conformation of an ATP-bound DnaK mutant (PDB ID: 4B9Q)  [69]  stabilized by introducing disulphide bridges at specific positions deduced from the structure of the ATP-Hsp110 homolog (Fig. 1 B ). The transition between the open and the closed structural states of Hsp70 is triggered by the ATP binding to the NBD which induces a conformational change of the inter-domain linker and the opening of the SBD  [61, 62] , through an allosteric mechanism which is not fully elucidated  [12, [61] [62] [63] [64] [65] [66] [67] [68].
3 A ) and the one calculated from the MD simulations of ADP-DnaK (Fig. 3 A ) show two peaks: one at 30 A ˚and a distinct shoulder at 60 A ˚, reflecting the bilobal shape of the protein in this nucleotide-binding state. The intermediate structure of ATP*-DnaK is different from the very recent crystallized triple mutant ATP-bound DnaK (E47C, T199A, F529C) in an open conformational state with a SBD-a covalently bound to the NBD through a disulphide bridge 47C-529C (PDB ID: 4B9Q)  [69]. In ATP*, as in ATPbound DnaK (Fig. 1 B ), the linker forms a b strand interacting with the loop between the strands b2 and b2' of the SBD-b. However, the position of the linker in ATP-bound DnaK is shifted relative to the one in ATP*: a contact is formed between D393 and I418 in ATP-bound structure whereas a contact is formed between D393 and N415 in ATP* (see next subsection)  [69].
Because of this limited sampling issue, only the FEPs which are similar in different trajectories in a given nucleotide-binding state were considered. Convergence of the FEPs. For each c i , the similarity index H(c i ) computed from two MD trajectories of DnaK in the same nucleotide-binding state of the NBD was calculated, i. e. H(c i ) APO1/APO2 , H(c i ) ADP1/ADP2 and H(c i ) ATP1/ATP2 (see Eq. 2). For the APO state, 471 of 597 FEPs were similar (''converged'') in the two MD trajectories representing 79% of all c i angles (defining the DATA for the APO state ; {c i } APO ). For the ADP and ATP bound states, 449 of 597 (DATA  ADP ; {c i } ADP ) and 447 of 597 (DATA ATP;{c i } ATP ) FEPs were similar (''converged''), respectively. The similar FEPs are located all along the amino-acid sequence, both in secondarystructures and in loops, and have various shapes from strongly harmonic FEPs to multiple-minima FEPs  [71].
Subset strongly, significantly and weakly influenced correspond to 1-H(c i ). 0. 7, 0. 3,1-H(c i )#0. 7 and 1-H(c i )#0. 3, respectively. doi:10. 1371/journal. pcbi. 1003379. t002 from i = 1 to 5. The most probable structure within each minimum A i of the MD run ATP1 is shown in Fig. S5 : the structure corresponding to the minimum A 1 is an elongated structure, without interactions between the NBD and the SBD and closely similar to the initial one (PDB ID: 2KHO)  [70]. In the basin corresponding to the minimum A 2 , the SBD rotates around the linker and get close to the NBD but without strong interactions between the two main domains. In the structure of the global minimum A 3 of the MD run ATP1, the SBD is docked to the lobe I of the NBD (Fig. S5 ). Note that the lid of the SBD (SBD-a) is docked in the subdomain B of the lobe I of the NBD.
A large comparison between the residues elucidated from the analysis of the FEPs similarity computed from the MD simulations (Table  3 ) and the experimental database of key residues relevant for the communication in DnaK available in the literature, shows that most of the residues identified by our methodology have been found to be experimentally relevant for the allosteric communication in Hsp70 s and that new residues can be proposed to play a role according to the present MD calculations. In more detail, the comparison in Table  3  can be summarized as follows. In the NBD, mutation of residue T13 in bovine Hsc70 (T11 in E. coli DnaK) to S13 completely abrogated the communication between the NBD and the SBD  [95]. Residue T13 was indeed found from the analysis of the FEP of c 12 , which belongs to the nucleotide-binding pocket (Fig.
MD simulations All unbiased all-atom MD simulations in explicit water of E. coli DnaK using the NMR-derived structure (PDB ID: 2KHO in which the missing atoms of ARG and TYR were added)  [70]  were carried out with the GROMACS software package  [105]  and the GROMOS96 ffG43a1 force field  [106, 107]. The time step used in all simulations was 0. 001 ps and the list of neighbors was updated every 0. 005 ps with the 'grid' method and a cut-off radius of 1 nm. The coordinates of all the atoms in the simulation box were saved every 2 ps. The initial velocities were chosen randomly. We used the NPT ensemble with a cubic box. The temperature and pressure were kept to the desired value by using the Berendsen method  [108]  and an isotropic coupling for the pressure (T = 300 K, t T = 0. 1 ps; P0 = 1 bar, coupling time t P = 0.
S10 A ). To reduce the computational cost, the APO and ADP-bound DnaK MD runs were stopped when the RMSD was converged to a constant value with a standard deviation lower than 0. 5 A ˚(Fig. S10 B ) during at least 50 ns, resulting in six MD runs of different durations. The 2 MD runs of ATP-DnaK were continued after convergence of the RMSD up to 0. 5 ms in order to explore a possible opening of the SBD which did not occur however on that time-scale. MD runs of APO-Hsp70. The initial structure of the nucleotide-free DnaK (PDB ID: 2KHO)  [70]  was solvated in a cubic box with 136508 SPC water molecules keeping a minimum distance of 0. 9 nm between the solute and each face of the box. We used periodic boundary conditions with an initial value of the box side of 16. 112 nm. The charge of the system was neutralized by adding 25 Na + counter-ions.
The energy of the system was first optimized with the ''steepest descent minimization'' algorithm and then by using a ''conjugate gradient'' algorithm. We performed two runs with different initial conditions. The total simulation time was 151 ns for the MD run APO1 and 360 ns for the MD run APO2. MD runs of ADP-Hsp70. The topology files of the ADP and ATP molecules were constructed with the PRODRG program (version 2. 5 BETA)  [110]. We added one ADP molecule and one divalent ion (Ca 2+ ) to the nucleotide-free DnaK (PDB ID: 2KHO)  [70]. The initial coordinates of the ADP molecule were extracted from those of the ADP molecule in the X-ray structure of the isolated NBD of ADP-bound hHsp70 (PDB ID: 1HJO  [111] ) after Table  4. List of residues found to be relevant for conformational dynamics of human Hsp70 deduced from NMA of human Hsp70  [49, 92]  and from dPCA of E.
Location NMA dPCA NBD IA NBD IB L50-F78-K100-Y107-K108-E110-K112 K77-F78-G79-D80-D99-K100-P101-K102 NBD IIA R193 G215-I216 NBD IIB R247-H249-K250-K251-S254-Q255-R299 K250-K251-D252-I253 linker V388-Q389-D390-L391 SBD-b G408-V409-T429-Y431-S432-Q441-S462-I464-A467-P468-R469- G470-V471-P472-Q473-D492-K493-S494-T495-K497-A498-N499-K500 G408-V409-M410-T411-D433-N434-Q435-P436-G445-E446-R447-A448- P465-P466-A467-A468-R469-K493-S494-T495-G496 SBD-a Y525-K526-Q532-R533-R535-Y545-K559-K561-S563-E564-A565-D566-K567- K568-K569-K573-W580-E588-K589-D590-E591-F592-H594-R596-K597-Q601 N505-D506-K507-G508-E521-A522-E523-K524 Residues common to the two analyses are in bold. Residues of hHsp70 which are not identical in E. coli Hsp70 are in italic font. doi:10. 1371/journal. pcbi. 1003379. t004 alignment of the backbone atoms of the residues 5 to 377 of the hHsp70 NBD on those of the DnaK NBD (PDB ID: 2KHO) by using the VMD software  [112].
The protein was solvated with 136488 SPC water molecules  [113]  in a cubic box of the same dimensions as in the MD runs APO (see above), and 26 Na + counterions were added to neutralize the system. The ADP-bound DnaK initial structures were relaxed during 600 ps with the protein structure restrained to the experimental structure (PDB ID: 2KHO) and with no constraints on the solvent and on the ligands (Dataset S1 in supporting information). We performed two MD runs ADP1 and ADP2 with different initial conditions. The total simulation time was 164 ns for the MD run ADP1 and 151 ns for the MD run ADP2. MD runs of ATP-Hsp70. The ATP-Hsp70 structure was built using the NMR-derived structure of E. coli DnaK (PDB ID: 2KHO)  [70]  with ADP and one divalent ion inserted as described above. The initial coordinates of the ATP molecule was built by adding a phosphate group to the ADP molecule in the free-space available within the NBD of the ADP-bound DnaK structure.
The precise initial location of the third phosphate group was not crucial because the ATP-bound DnaK structure was relaxed during 600 ps with the protein structure restrained to the experimental structure (PDB ID: 2KHO) and with no constraints on the solvent and on the ligands (Dataset S2 in supporting information). We compared the ATP-bound construct relaxed after 600 ps with available X-ray structures of isolated NBD of hHsp70 bound to ADP and PO 4 (PDB IDs: 1S3X  [114] , 3AY9  [115] , 3JXU  [116] ). The relaxation of the ATP molecule using the present force-field displaced the added phosphate group to a position close to the experimental position of the PO 4 group in the isolated NBD of hHsp70. Comparison with the new published X-ray full-length ATP-bound open structure (PDB ID: 4B9Q)  [69] , confirmed that the initial relaxed position of the ATP molecule (which mimic the ATP binding position in the full-length E.
B and C and Fig. S2 C): T11, G197, G198, T199, I271 and G341. The contacts depicted in Fig. 4 for the intermediate state ATP*-DnaK can be compared qualitatively with those observed in the very recent crystallized triple mutant ATP-bound DnaK [69]. Although the structure of the intermediate state found in MD and the crystallized open conformation of DnaK are quite different as discussed above, many contacts between ATP and the NBD found in ATP*-DnaK (Fig. 4 B and C) are also found in the experimental structure of ATP-bound DnaK (PDB ID: 4B9Q) [69]. Except D195-ATP, at a contact distance (see Methods section) of 4. 9 A ˚in.
Some processes have already been described earlier (see for example  [5, 8] ); they will be summarized here in order to present a complete overview of the data treatment. In particular most of the in-flight procedures used to obtain information on the detector and AOTF (Acousto Optical Tunable Filter) characteristics have been analysed and discussed in great detail in  [8, 9]  and only the results and conclusions will be of interest here. Improvements made since then will be described and commented. New developments will be explained in full detail. The format and content of the archived data (PSA Levels 2 and 3) on the ESA (European Space Agency) servers will be clarified. The SOIR archive is at version v4. 0 at the time of submission of this text.
Optional different binning options can be defined and have been used. For completeness they are also given in Table  2. The spectral sampling interval varies from 0. 029 to 0. 032 cm -1 in diffraction order 101 to 0. 052 to 0. 06 cm -1 in order 194, increasing with the pixel number and the diffraction order. In the following, we will describe each step of the data pipeline from raw (PSA Level 2) to calibrated (PSA Level 3) data. We will also describe some ancillary information useful for the analysis and interpretation of the SOIR spectra.
These temperature changes are partially measured by the thermal sensors inside the SOIR instrument, but are difficult to correlate with the observed variations of the spectral calibration. To overcome this problem, we implemented a second calculation step in the spectral calibration on each spectrum, based on the presence of absorption lines with well -known positions in spectral databases such as HITRAN  [14]. Pressure-induced line shifts were neglected since the resolution and the spectral sampling of SOIR are larger than the pressure shifts induced by collisions. The polynomial degree is five at maximum, though third degree polynomials are used in most of the cases. For some SOIR diffraction orders where few absorption lines are present, the degree is set to lower values, depending on the number of spectral lines available for the calibration.
The calibration results from the comparison of the observed positions of the selected lines and the positions found in spectroscopic databases. A spectral error is calculated for each spectrum by comparing the position of selected atmospheric lines in HITRAN and in the calibrated SOIR spectra. It varies from 0. 005 to 0. 02 cm -1. This calibration procedure will be illustrated by the following example: the order 190 is considered, which corresponds to the 4246. 15 -4282. 62 cm -1 spectral interval. In this range, CO is the main absorber, although absorption due to HF and HDO is also present. One band of CO is observed in the spectrum: the (2-0) band of the main isotopologue 12 C 16 O (CO26, using the HITRAN notation). Figure  4  illustrates the procedure on one such spectrum. In the top Panel, the observed spectrum is compared to the simulated one obtained by using the previous pixel-wavenumber relation whereas in the bottom Panel the calibration is based on the position of the CO selected lines, indicated with stars in the top Panel.
The automatic procedure detects when the calibration is starting to deteriorate (either the number of usable lines is too low, or the regression between observed and HITRAN positions is too poor). Calibration for such spectra is then the same as the one determined for the closest spectrum for which the calibration was deemed acceptable. The direct calibration F(p) is reported in the archive at PSA level 3 in the DATA directory together with the spectra. The inverse relation is not given since it can easily be calculated from the direct one.
Description of the archived data The SOIR PSA archive can be found on the ESA servers at the address http://www. rssd. esa. int/index. php?project=PSA&page=vex. This paper describes the Version 4. 0 of the SOIR archive, which corresponds to Release 3 and Revision 0. The format has changed significantly compared to previous versions. The observations are separated in data sets at PSA Levels 2 and 3 as a function of the mission extension during which they were recorded: the nominal mission covers observations up to orbit 530, the first extension up to orbit 1136, the second extension up to orbit 1584, the third extension up to orbit 2080 and the fourth one for the latest orbits. The name of an observation is based on the date it was recorded and its type. Because the orbital period of VEX is 24 hours, different measurements can be done during one day, for example an ingress followed by an egress.
The format is YYYYMMDD_TCC where YYYY is the year of the orbit, MM is the month, DD is the day, T is the type of measurement and CC is the measurement number during that orbit, starting from 1. The types of observations are summarized in Table  4. Besides egresses and ingresses, which correspond to solar occultation observations carried out when exiting or entering penumbra, there are also Sun and atmospheric full scans (all orders are scanned sequentially), mini-scans (the AOTF frequency is scanned over a short interval) and nadir (looking at the surface of the planet). Only ingresses and egresses are given in the archive at PSA Level 3. We will briefly describe the files making up the Level 3 PSA archive, with special emphasis on the calibration (CALIB directory) and spectra files (DATA directory).
Indeed the complete archive contains a lot of files describing the formats, the content, retracing the history of the archive. They can be found under the DOCUMENT and CATALOG folders. The archive BROWSE directory contains spectra Figs. in JPG format: the sum of all pixels as a function of time at PSA Level 2, and all the transmittance spectra at PSA Level 3. We will only explain how the different calibrations and the instrument characterization are implemented in the archive. All the ASCII data are given in TAB files. They are accompanied by LBL files, which describe the content of the data files. All calibration files can be found under the CALIB directory at PSA Levels 2 and 3. Several files can be found concerning: (1) The resolution file (RESOL_BINNINGXX. TAB, with XX the binning option) contains the values of the resolution (FWHM of a Gaussian ILS) for each diffraction order and bin; (2) The AOTF tuning function file (AOTF_F_WN.
Conclusions We have presented several improvements in the calibration and characterization of the SOIR instrument introduced in the data pipeline since  [8]. They concern mainly the spectral calibration which has been improved by not only considering solar lines but also atmospheric absorption lines in each individual order of the instrument. The characterization of the ILS has also improved by considering a large number of atmospheric lines compared to the limited number of solar lines that were previously used for this purpose. The detailed investigation of the noise on each individual pixel has led to the determination of noise spectra defined for each spectrum, as it was shown that this noise pattern was highly dependent on the observation settings. We have also described the data archive for the SOIR instrument, describing what information should be found in which file.
Sampling The study was carried out in the locality of l'Escala in the Northwest Mediterranean sea (42°06′52″N, 3°10′07″E). All samples were collected between 10 and 14 m depth from a dense Dysidea avara population, sitting on a rocky wall facing NW. The temporal variability in production of avarol and its monoacetylated derivative was assessed by sampling randomly ten Dysidea avara individuals once a month over almost two years (from March 2009 to January 2011). Temperatures were continuously recorded in situ by a Stowaway Tidbits ® , autonomous data logger (0. 2 °C precision, 0. 15 °C resolution), placed at the study site, at 14 m of depth. The end of the reproductive period was assessed by verifying the presence of brooded embryos in the sponge tissue through a stereomicroscope. Histological observations by the authors revealed that the reproductive period (i.