The overall fold of <i>H</i>. <i>pylori</i> PseH.

<p>(A) Stereo diagram of the structure of the PseH monomer. β-strands and α-helices are represented as arrows and coils and each element of the secondary structure is labeled and numbered as in text. The bound AcCoA molecule is shown in black. (B) The topology of secondary structure elements PseH. The α-helices are represented by rods and β-strands by arrows. Residue numbers are indicated at the start and end of each secondary structure element. (C) The molecular surface representation of PseH showing the AcCoA-binding tunnel between strands β4 and β5, which is a signature of the GNAT fold.</p

Refinement statistics.

<p>^a</p><p></p><p></p><p>R=</p><p></p><p></p><p></p><p>∑h</p><p>|</p><p>(</p><p>|</p><p>Fobs</p>|<p></p>−<p>|</p><p>Fcalc</p>|<p></p>)<p></p>|<p></p><p></p><p></p><p></p><p></p><p>∑h</p><p>|</p><p>Fobs</p>|<p></p><p></p><p></p><p></p><p></p><p></p><p></p><p></p><p>^b The free R-factor was calculated on 5% of the data omitted at random.</p><p>Refinement statistics.</p

The stereoview of the electron density for AcCoA bound in the active site of PseH.

<p>The cofactor molecule is shown in CPK representation and coloured according to atom type, with carbon atoms in orange, nitrogen in blue, oxygen in red, phosphorus in magenta and sulphur in yellow. Only the protein residues that form hydrogen bonds or van der Waals contacts with the cofactor molecule are shown for clarity. Protein carbon atoms are coloured black. The hydrogen bonds important for recognition of the cofactor are shown. The map was calculated at 2.3 Å resolution with coefficients |F_obs| − |F_calc| and phases from the final refined model with the coordinates of AcCoA deleted prior to one round of refinement. The map is contoured at 3.0-σ level.</p

The CMP-pseudaminic acid biosynthesis pathway in <i>H</i>. <i>pylori</i> [10].

<p>The CMP-pseudaminic acid biosynthesis pathway in <i>H</i>. <i>pylori</i> [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0115634#pone.0115634.ref010" target="_blank">10</a>].</p

Interactions between the docked substrate UDP-4-amino-4,6-dideoxy-β-<i>L</i>-AltNAc, acetyl moiety of the cofactor and protein residues in the active site of PseH in the modeled Michaelis complex.

<p>The protein backbone is shown as ribbon structure in light grey for clarity of illustration. The substrate and AcCoA molecules are shown in ball-and-stick CPK representation and coloured according to atom type, with carbon atoms in black, nitrogen in blue, oxygen in red, phosphorus in magenta and sulphur in yellow. Only the protein side-chains that interact with the substrate are shown for clarity. The C4-N4 bond of the substrate (labeled) is positioned optimally for the direct nucleophilic attack on the thioester acetate, with the angle formed between the C4 of the amino-altrose, N4 of amino-altrose and the thioester carbonyl carbon being approximately 120°. The water molecule that is hydrogen bonded to the side-chains of Ser78 and Thr80, and is located within a hydrogen-bond distance of the 3’-hydroxyl of the modeled 4’-amino-altrose, is represented as a grey-blue ball. Deprotonation of the substrate’s amine group may occur via the 3’-hydroxyl of the altrose and this intervening water molecule.</p

The structural similarity between the nucleotide-binding pocket in MccE and the putative nucleotide-binding site in PseH.

<p>The positions of the protein side-chains that form similar interactions with the nucleotide moiety of the substrate and with AcCoA are shown in a stick representation. The 3'-phosphate AMP moiety of CoA is omitted for clarity. (A) Key interactions between the protein and the nucleotide in the complex of the acetyltransferase domain of MccE with AcCoA and AMP. The protein backbone is shown as ribbon structure in light green for clarity of illustration. The AMP and AcCoA molecules are shown in ball-and-stick CPK representation and coloured according to atom type, with carbon atoms in black, nitrogen in blue, oxygen in red, phosphorus in magenta and sulphur in yellow. (B) The corresponding active-site residues in PseH and the docked model for the substrate UDP-4-amino-4,6-dideoxy-β-<i>L</i>-AltNAc. The protein backbone is shown as ribbon structure in light grey for clarity of illustration. AcCoA and modeled UDP-sugar are shown in ball-and-stick CPK representation and coloured according to atom type, with carbon atoms in black, nitrogen in blue, oxygen in red, phosphorus in magenta and sulphur in yellow.</p

X-ray data collection and processing statistics.

<p>Values in parentheses are for the highest resolution shell.</p><p>^a</p><p></p><p></p><p></p><p>Rmerge</p>=<p></p><p></p><p>(</p><p></p><p>∑h</p><p></p><p>∑i</p><p>|</p><p>I</p><p>hi</p><p></p>−<p>〈</p><p>Ih</p>〉<p></p>|<p></p><p></p><p></p>)<p></p><p></p><p></p><p></p><p>∑h</p><p></p><p>∑i</p><p>|</p><p>I</p><p>hi</p><p></p>|<p></p><p></p><p></p><p></p><p></p><p></p><p></p><p></p>, where <i>I_hi</i> is the intensity of the <i>i</i>th observation of reflection <i>h</i>.<p></p><p>X-ray data collection and processing statistics.</p

Structure–Activity Relationship for Sulfonamide Inhibition of Helicobacter pylori α‑Carbonic Anhydrase

α-Carbonic anhydrase of Helicobacter pylori (HpαCA) plays an important role in the acclimation of this oncobacterium to the acidic pH of the stomach. Sulfonamide inhibitors of HpαCA possess anti-H. pylori activity. The crystal structures of complexes of HpαCA with a family of acetazolamide-related sulfonamides have been determined. Analysis of the structures revealed that the mode of sulfonamide binding correlates well with their inhibitory activities. In addition, comparisons with the corresponding inhibitor complexes of human carbonic anhydrase II (HCAII) indicated that HpαCA possesses an additional, alternative binding site for sulfonamides that is not present in HCAII. Furthermore, the hydrophobic pocket in HCAII that stabilizes the apolar moiety of sulfonamide inhibitors is replaced with a more open, hydrophilic pocket in HpαCA. Thus, our analysis identified major structural features can be exploited in the design of selective and more potent inhibitors of HpαCA that may lead to novel antimicrobials

Structural comparison between the MZA and AAZ complexes of HpαCA.

<p>The MZA molecule is shown in all-atom representation with carbon atoms coloured in orange. The (mFo-DFc) sigmaA-weighted electron density for MZA is shown in green. The map was calculated at 2.2-Å resolution and contoured at 3.0-σ level. The AAZ molecule is shown in cyan. Protein surface is shown and coloured according to the electrostatic potential. The subtle rotation and shift of MZA with respect to AAZ does not break the hydrogen bond between the O3 atom of the carbonamide moiety and the side chain of Asn108. The weaker binding of MZA in comparison to AAZ is likely due to energetically unfavourable interaction between the additional aliphatic methyl group of MZA and partially negatively charged carbonyl oxygens of the main-chain peptides of Ala192 (C-O distance 4.3 Å) and Pro193 (C-O distance 3.5 Å).</p

Structure-based sequence alignment of HpαCA, NgCA, SspCA, TaCA and HCAII.

<p>The elements of the secondary structure and the sequence numbering for HpαCA are shown above the alignment. Conserved residues are highlighted in red. The conserved histidine residues coordinating the zinc ion are marked by an asterisk. The proton shuttle residue of HCAII (His64) is marked with an open circle. The corresponding residue in HpαCA is His85. The conserved residues that form a hydrophobic pocket in the active site are labeled with filled squares. The location of the catalytically important HCAII residue Thr199 which forms a hydrogen bond to the zinc-bound water (Wat263, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0127149#pone.0127149.g005" target="_blank">Fig 5</a>), thereby orienting its two lone electron pairs toward the two neighbouring water molecules (Wat318 and Wat338) in the active site is shown by an open square. The corresponding residue in HpαCA is T191.</p