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3nqh

    Table of contents
    1. 1. Protein Summary
    2. 2. Ligand Summary

    Title Crystal structure of a glycosyl hydrolase (BT_2959) from BACTEROIDES THETAIOTAOMICRON VPI-5482 at 2.11 A resolution. To be published
    Site JCSG
    PDB Id 3nqh Target Id 396210
    Molecular Characteristics
    Source Bacteroides thetaiotaomicron vpi-5482
    Alias Ids TPS25885,NP_811871.1, _0110.003913_, 332862 Molecular Weight 49621.71 Da.
    Residues 440 Isoelectric Point 6.56
    Sequence rktekvvnngipwfddrgeivnahgaciveengryylfgeyksdksnafpgfscyssddlvnwkfervv lpmqssgilgpdrvgervkvmkcpstgeyvmymhaddmnykdphigyatcstiageyklhgpllyegkp irrwdmgtyqdtdgtgylllhggivyrlskdyrtaeekvvsgvggshgespamfkkdgtyfflfsnlts wekndnfyftapsvkgpwtrqglfapegsltynsqttfvfplkcgedtipmfmgdrwsyphqasaatyv wmpmqvdgtklsipeywpswdvdklkpvnplrkgktvdlkkitfskeadwkveegrissnvkgstlsip ftgscvavmgetnchsgyarmnildkkgekiysslvdfyskandhatrfktpqlaegeytlvievtgis ptwtdktkriygsddcfvtitdivkl
      BLAST   FFAS

    Structure Determination
    Method XRAY Chains 1
    Resolution (Å) 2.11 Rfree 0.238
    Matthews' coefficent 2.58 Rfactor 0.195
    Waters 239 Solvent Content 52.33

    Ligand Information
    Ligands
    Metals

    Jmol

     
    Google Scholar output for 3nqh
    1. The structure and function of an arabinan-specific _-1, 2-arabinofuranosidase identified from screening the activities of bacterial GH43 glycoside hydrolases
    A Cartmell, LS McKee, MJ Pea, J Larsbrink - Journal of Biological , 2011 - ASBMB
     
    2. Crystal structure of 1, 3Gal43A, an exo-_-1, 3-Galactanase from Clostridium thermocellum
    D Jiang, J Fan, X Wang, Y Zhao, B Huang, J Liu - Journal of Structural , 2012 - Elsevier
     

    Protein Summary

    The carbohydrate binding module in our protein and its closest (but still very distant - 13% seq id) homolog 3c7e has a different position in respect to the central catalytic domain. You could see it by comparing the rigid http://fatcat.sanfordburnham.org/fatcat-cgi/cgi/fatcatextract.pl?-func=jmol&-base=51053&-sysdir=result&-file=3nqh_.3c7e_ and flexible http://fatcat.sanfordburnham.org/fatcat-cgi/cgi/fatcatextract.pl?-func=jmol&-base=51052&-sysdir=result&-file=3nqh_.3c7e_FATCAT alignments. You can see it on the simple move http://fatcat.sanfordburnham.org/fatcat-cgi/cgi/viewMorph.pl?-sysdir=result&-base=51052&-pdb1=3nqh_.pdb&-pdb2=3c7e_.pdb&-aln=3nqh_.3c7e_.txt&-file=3nqh_.3c7e_. Of course the movie is completely ridiculous, as the CBM goes through a "flat" (i.e. completely unphysical) conformation. Its sort of hilarious. I found this looking for a good example of FATCAT use and only after a while I realized this it our protein

    Only few of the 5-blade propellers of the GH43 family have already been deposited in the Protein Data Bank (3c2u, 2exh(i,j,k), 3c7(E,F,G,H,O and 3kst). The most similar regarding the overall fold and the active site residues is 3c7e(f,g,h,o) pdb structures, cocrystallized with sugar substrates(1). The novelty of the PX9010D target is that it is assembled of 2 domains, the catalytic propeller and beta-sandwich made of 5-beta strands on each side and a characteristic extended loop at the very C-terminal portion of the whole molecule. These loop residues 434-448 serve as a 'lid' covering the entrance into the catalytic side around the central tunnel of the propeller. If taking the homology with 3c7e arabinofuranohydrolase into account, the C-terminal beta-sandwich domain would represent a carbohydrate binding module with rmsd of 2.6A (DALI score 10.6). Overall fold is attached below in Fig.1.

    Overview of the overlapping catalytic residues of the 3c7g PDB entry (in magenda) and its sugar ligand (green) over PX9010D (white residues and cyan ribbon), pictures the completely conserved general acid GLU207 residue, ASP282 or GLU60 possibly acting as general base, and ASP163 functioning as a pKa modulating and substrate orienting residue. 3c7g has no lid covering the 5-fold tunnel entrance, which exists in PX9010D(Fig2.).

    The beta-sandwich domain is not connected through a clean velcro mechanism with the propeller (as seen in many other similar structures), instead a short 1 turn helix (in yellow) connection meets the velcro point on the side of the beta-sandwich domain (yasara hydrogen bond network picture

    PX9010D was cocrystallized with glycerol, which occur to bind in the active pocket, mimicking well the positions of the hydroxy groups with the ones from superposed sugar substrates from homolog structures(3c7e,e,f,g,h,o).

    A DALI search of the whole molecule revealed absolute conservation of TRP83, GLU105, PHE219, TYR175. Part of the catalytic residues as suggested by nearest sequence homology include GLU207, ASP163, ASP282. All the conserved residues are located in the very positively charged entrance tunnel of the propeller domain.

     

    The positive pole of the dipole moment originates in the channel around the axis of the propeller and the CBM domain and points towards the C-terminal beta-sheet domain, in the direction of the lid loop, when looking from the side (Fig4). The channel around the five-fold propeller axis is known to be the binding pocket for sugar substrates which are being hydrolyzed by the system. Complete conservation of three catalytic acidic residues in that pocket including partial conservation of aromatic residues around the active side as compared to 2EXH,2EXJ and 3C7G,E,F,H,O, implicates that stacking interactions and general base/acid mechanism interactions are responsible for the hydrolysis. The initial binding of the substrates though, could result from an electrostatic force, in particular since the tunnel is highly acidic and the lid contains 'RKTK' positively charged finger-shaped surface covering that acidic groove. This positive charged loop possibly restricts the length of xylose units that can enter the active site. When overlapping 2EXH inverting glycoside hydrolase with its XYS substrate on PX9010D, the lid loop sits right above the substrate, which in our structure was substituted by a GOL molecule overlapping one xylopyranosyl unit from XYS.

    Since the 3 catalytic residues are positioned in a plane perpendicular to the tunnel axis where the sugars are docked for the hydrolysis, the last to be separated unit seems (at least on few structures with longer substrates) to be positioned in or above that plane, thus forming a kink on this unit along the oligomeric substarte. If the dipole moment of PX9010D pointing outwards the central propeller tunnel has any meaning, it could indicate the direction of 'extraction' of the separated sugar unit. It is possible, that type of residues spanning that tunnel governs the extraction speed/timing.

    Ligand Summary

    Reviews

    References

     

    No references found.

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    Files (5)

    FileSizeDateAttached by 
    CATALYTICside3c7g.jpg
    Catalytic residues of PX9010D in white conserved with the ones from (3c7g) arabinofuranohydrolase(magenda and green)
    241.59 kB19:04, 14 Apr 2010cbtrameActions
     DIPOL-TOP.jpg
    PX9010D Dipole Moment as calculated by http://bioinfo.weizmann.ac.il/dipol-bin/dipol2j.cgi
    356.91 kB01:21, 15 Apr 2010cbtrameActions
     ELECTRO-TUNNEL-LID-DELPHI.png
    Delphi electrostatic surface near the catalytic pocket located below the PX9010D lid
    738.54 kB01:23, 15 Apr 2010cbtrameActions
    FOLD-chim1.jpg
    Overall PX9010D fold: catalytic 5-blade beta-propeller domain with C-terminal beta-sandwich CBM module and its "LID" loop directing the intrance into the catalytic tunnel
    337.88 kB19:08, 14 Apr 2010cbtrameActions
    YASA-thinTOP-OVERALL.png
    Yasara representation of the hydrogen bond network in PX9010D
    484.13 kB00:12, 15 Apr 2010cbtrameActions
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