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The Open Protein Structure Annotation Network
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3kkf

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

    Title Crystal structure of Putative antibiotic biosynthesis monooxygenase (NP_810307.1) from Bacteriodes thetaiotaomicron VPI-5482 at 1.30 A resolution. To be published
    Site JCSG
    PDB Id 3kkf Target Id 396203
    Molecular Characteristics
    Source Bacteroides thetaiotaomicron vpi-5482
    Alias Ids TPS26658,NP_810307.1, 3.30.70.900, 326096 Molecular Weight 11999.34 Da.
    Residues 104 Isoelectric Point 5.19
    Sequence aennmvrlsriiidperleeynaylkeeievsmrlepgvlvlyavaekerpnhvtileiyadeaayksh iatphfkkykegtldmvqmlelidatplipglkmk
      BLAST   FFAS

    Structure Determination
    Method XRAY Chains 1
    Resolution (Å) 1.30 Rfree 0.167
    Matthews' coefficent 1.88 Rfactor 0.135
    Waters 147 Solvent Content 34.52

    Ligand Information
    Ligands
    Metals

    Jmol

     
    Google Scholar output for 3kkf

    Protein Summary

    Protein BT_1394 (JCSG target ID 396203, JCSG target accession code PX8150A, GenBank accession code NP_810307.1) is a 131-amino acid long protein from Bacteriodes thetaiotaomicron, an anaerobic, gram-negative bacterium that is prevalent as part of the human gut microbiome.  The protein is annotated as being a putative flavoredoxin and as belonging to the antibiotic biosynthesis monooxygenase (ABM) protein family (Pfam PF03992, clan CL0032).

    The structure of an N-terminally truncated version of NP_810307.1 (residues 28-131) was solved by the Se-Met single-wavelength anomalous dispersion (SAD) method to a resolution of 1.3 Angstroms.  The structure of a monomer, which represents the content of an asymmetric unit, adopts an alpha-beta fold that fastSCOP classifies as belonging to the dimeric alpha+beta barrel superfamily in the SCOP database (Figure 1a).

    Proteins in this superfamily have been found to dimerize via their beta-sheet to form a core beta-barrel that is flanked by helices, and NP_810307.1 is no exception to this (Figure 1b).  In addition to the crystal packing evidence, analysis of the NP_810307.1 structure by the PISA server supports this dimeric form as being a stable assembly.

    Figure 1.  (a) Structure of a monomer of NP_810307.1 gradiently colored from the N- (blue) to the C-terminus (red).  (b)  Structure of NP_810307.1 in its dimeric form, its most probable physiologically relevant oligomeric state (monomers A and B are represented in yellow and blue ribbon, respectively; calcium ions are represented as magenta spheres; and acetate ions are represented as green sticks).

    (a)                                                                                (b)

    PX8150A-gradient.png      PX8150A-dimer.png

     

    Searches by DALI and SSM yield many structural homologs of NP_810307.1 (Tables 1 and 2), several of which are also JCSG structures (e.g., PDB codes: 3bm7, 3e8o, 2qyc)

    Table 1.  Structural homologs of NP_810307.1 as assessed by DALI.

    N PDB Z-score RMSD LALI NRES %ID TITLE
    1 2omo 15.8 1.5 98 99 15 DUF176
    2 2gff 15.8 1.5 96 97 22 LSRG PROTEIN
    3 3bm7 15.6 1.6 97 106 19 PROTEIN OF UNKNOWN FUNCTION WITH FERREDOXIN-LIKE (3bm7 TOPSAN)
    4 1x7v 15.1 1.8 98 98 19 PA3566 PROTEIN (Sanders et al. 2005)
    5 1y0h 14.4 1.7 97 98 16 HYPOTHETICAL PROTEIN RV0793 (Lemieux et al. 2005)
    6 2fb0 13.6 1.9 94 94 17 CONSERVED HYPOTHETICAL PROTEIN
    7 2bbe 13.6 1.8 97 103 21 HYPOTHETICAL PROTEIN SO0527
    8 1tuv 13.6 1.7 93 103 17 PROTEIN YGIN (Adams et al. 2005)
    9 1r6y 13.5 1.7 93 103 17 PROTEIN YGIN (Adams et al. 2005)
    10 3e8o 13.1 2.5 99 102 15 UNCHARACTERIZED PROTEIN WITH ERREDOXIN-LIKE FOLD (3e8o TOPSAN)

     

    Table 2.  Structural homologs of NP_810307.1 as assessed by SSM.

    N PDB Q-score RMSD TITLE
    1 2omo 0.7526 1.262 PUTATIVE ANTIBIOTIC BIOSYNTHESIS MONOOXYGENASE FROM NITROSOMONAS EUROPAEA
    2 1x7v 0.7461 1.343 PA3566 FROM PSEUDOMONAS AERUGINOSA (Sanders et al. 2005)
    3 2gff 0.7328 1.372 YERSINIA PESTIS LSRG
    4 1y0h 0.6927 1.624 STRUCTURE OF RV0793 FROM MYCOBACTERIUM TUBERCULOSIS (Lemieux et al. 2005)
    5 2fb0 0.6638 1.541 CONSERVED PROTEIN OF UNKNOWN FUNCTION FROM BACTEROIDES THETAIOTAOMICRON VPI-5482 AT 2.10 A RESOLUTION, POSSIBLE OXIDOREDUCTASE
    6 3bm7 0.648 1.504 PROTEIN OF UNKNOWN FUNCTION WITH FERREDOXIN-LIKE FOLD (NP_420935.1) FROM CAULOBACTER CRESCENTUS AT 1.35 A RESOLUTION (3bm7 TOPSAN)
    7 1r6y 0.6085 1.604 YGIN FROM ESCHERICHIA COLI (Adams et al. 2005)
    8 1tuv 0.6062 1.619 YGIN IN COMPLEX WITH MENADIONE (Adams et al. 2005)
    9 2qyc 0.5961 1.859 DIMERIC FERREDOXIN-LIKE PROTEIN (NP_888056.1) FROM BORDETELLA BRONCHISEPTICA AT 1.90 A RESOLUTION (2qyc TOPSAN)
    10 2pd1 0.5914 2.187 NE2512 PROTEIN OF UNKNOWN FUNCTION FROM NITROSOMONAS EUROPAEA

     

    A superposition of NP_810307.1 with several of the structural homologs from DALI and SSM (2fb0, 1tuv, 1y0h, and 3e8o) is shown in Figure 2a and reveals very close structural similarity.  An interesting difference involves the formation of the dimer.  In the case of NP_810307.1 and structural homologs 2fb0, 1tuv, 1y0h, and 3e8o, the structures are true homodimers (i.e., the dimer is formed by the association of two individual monomers), whereas in the case of 3f44 (one of the top FFAS hits), the structure is a pseudo-dimer in that it is formed by a domain repeat in a single polypeptide chain (see 3f44 TOPSAN).

    Figure 2.  (a) Superposition of NP_810307.1 (yellow) with PDB codes: 3f44 (cyan), 2fb0 (blue), 1tuv (green), 1y0h (red), and 3e8o (magenta).  Circled in red is the putative active site.  (b)  A view of the superimposed putative active sites showing that in this and many of its structural homologs, a ligand(s) is bound at this site. These ligands include acetate in the case of NP_810307.1, 2fb0, and 1y0h, an unidentified ligand (UNL) in 3e8o, and menadione in 1tuv.  Coloring scheme is the same as in (a).

    (a)                                                                                    (b)

    PX8150A-superpose-overall-mod.png    PX8150A-superpose-activesite.png

    Ligand Summary

    In NP_810307.1 as well as in a number of its structural homologs, a ligand(s) is bound at a similar corresponding location (Figures 2ab).  In the case of NP_810307.1, an acetate is bound at this location, as is the case with structural homologs 2fb0 and 1y0h.  In the case of 1tuv, menadione, which is a product of the reaction that this protein is speculated to catalyze, is bound at this site (Adams et al. 2005).

    A ConSurf analysis of residue conservation between NP_810307.1 and its top FFAS hits reveals that many of the residues surrounding this site are highly conserved (Figure 3).  These residues in NP_810307.1 include Glu-55, Ser-59, Glu-63, Leu-69, Ile-83, Glu-85, His-96, Thr-99, and His-101.

    Figure 3.  ConSurf analysis of the putative active site of NP_810307.1.  Acetate, as well as other adjacent solvent molecules (PEG and ethylene glycol (EDO)) are shown as green sticks.

     

    consurf_legend.png

    PX8150A-consurf-activesite.png

     

    An electrostatic surface calculation reveals that the base of the pocket in which the acetate is bound is negatively charged (Figure 4).

    Figure 4.  Electrostatic surface potential map of NP_810307.1.  The acetate ion in NP_810307.1 is represented as green sticks.

    PX8150A-emap.png

     

    The highly conserved nature of this pocket along with the fact that a number of structural homologs contain a ligand at this location strongly suggests that this could be the active site of NP_810307.1.

     

    Reference(s):

    Structural and biochemical evidence for an enzymatic quinone redox cycle in Escherichia coli: identification of a novel quinol monooxygenase.  Adams MA, Jia Z.  J Biol Chem. 2005 Mar 4;280(9):8358-63.

    The X-ray crystal structure of PA3566 from Pseudomonas aureginosa at 1.8 A resolution.  Sanders DA, Walker JR, Skarina T, Savchenko A. Proteins. 2005 Oct 1;61(1):209-12.

    The crystal structure of Rv0793, a hypothetical monooxygenase from M. tuberculosis.  Lemieux MJ, Ference C, Cherney MM, Wang M, Garen C, James MN. J Struct Funct Genomics . 2005 Dec;6(4):245-57.

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