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Structural basis of Lewis(b) antigen binding by the Helicobacter pylori adhesin BabA.

Hage N, Howard T, Phillips C, Brassington C, Overman R, Debreczeni J, Gellert P, Stolnik S, Winkler GS, Falcone FH - Sci Adv (2015)

Bottom Line: No conformational change occurs in BabA upon binding of Le(b), which is characterized by low affinity under acidic [K D (dissociation constant) of ~227 μM] and neutral (K D of ~252 μM) conditions.Binding is mediated by a network of hydrogen bonds between Le(b) Fuc1, GlcNAc3, Fuc4, and Gal5 residues and a total of eight BabA amino acids (C189, G191, N194, N206, D233, S234, S244, and T246) through both carbonyl backbone and side-chain interactions.The structural model was validated through the generation of two BabA variants containing N206A and combined D233A/S244A substitutions, which result in a reduction and complete loss of binding affinity to Le(b), respectively.

View Article: PubMed Central - PubMed

Affiliation: School of Pharmacy, University of Nottingham, University Park, Nottingham NG7 2RD, UK.

ABSTRACT
Helicobacter pylori is a leading cause of peptic ulceration and gastric cancer worldwide. To achieve colonization of the stomach, this Gram-negative bacterium adheres to Lewis(b) (Le(b)) antigens in the gastric mucosa using its outer membrane protein BabA. Structural information for BabA has been elusive, and thus, its molecular mechanism for recognizing Le(b) antigens remains unknown. We present the crystal structure of the extracellular domain of BabA, from H. pylori strain J99, in the absence and presence of Le(b) at 2.0- and 2.1-Å resolutions, respectively. BabA is a predominantly α-helical molecule with a markedly kinked tertiary structure containing a single, shallow Le(b) binding site at its tip within a β-strand motif. No conformational change occurs in BabA upon binding of Le(b), which is characterized by low affinity under acidic [K D (dissociation constant) of ~227 μM] and neutral (K D of ~252 μM) conditions. Binding is mediated by a network of hydrogen bonds between Le(b) Fuc1, GlcNAc3, Fuc4, and Gal5 residues and a total of eight BabA amino acids (C189, G191, N194, N206, D233, S234, S244, and T246) through both carbonyl backbone and side-chain interactions. The structural model was validated through the generation of two BabA variants containing N206A and combined D233A/S244A substitutions, which result in a reduction and complete loss of binding affinity to Le(b), respectively. Knowledge of the molecular basis of Le(b) recognition by BabA provides a platform for the development of therapeutics targeted at inhibiting H. pylori adherence to the gastric mucosa.

No MeSH data available.


Related in: MedlinePlus

Interactions between BabA amino acids and Leb residues.(A) Fuc1 forms hydrogen bonds with the carbonyl backbone of C189, G191, and N194 and the side chain of T246. (B) Fuc4 forms a water-mediated hydrogen bond with the N206 side chain. (C) GlcNAc3 forms hydrogen bonds with the D233 and S244 side chains. (D) Gal5 forms hydrogen bonds with the carbonyl backbone and side chain of S244. Gal5 also forms hydrogen bonds with a water molecule structured by the side chains of D233 and S234. Fucose, galactose, and N-acetylglucosamine residues are colored orange, yellow, and blue, respectively. Hydrogen bonds are represented by dotted black lines.
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Figure 3: Interactions between BabA amino acids and Leb residues.(A) Fuc1 forms hydrogen bonds with the carbonyl backbone of C189, G191, and N194 and the side chain of T246. (B) Fuc4 forms a water-mediated hydrogen bond with the N206 side chain. (C) GlcNAc3 forms hydrogen bonds with the D233 and S244 side chains. (D) Gal5 forms hydrogen bonds with the carbonyl backbone and side chain of S244. Gal5 also forms hydrogen bonds with a water molecule structured by the side chains of D233 and S234. Fucose, galactose, and N-acetylglucosamine residues are colored orange, yellow, and blue, respectively. Hydrogen bonds are represented by dotted black lines.

Mentions: The crystallographic model indicated that binding is mediated by a network of hydrogen bonds between Leb Fuc1, GlcNAc3, Fuc4, and Gal5 residues and a total of eight BabA amino acids. Fuc1 forms hydrogen bonds with the carbonyl backbone groups of C189, G191, and N194, and the hydroxyl group of the T246 side chain (Fig. 3A). Fuc4 interacts with the hydroxyl group of the N206 side chain through a water-mediated hydrogen bond (Fig. 3B). The GlcNAc3 residue forms two hydrogen bonds with the carboxyl and hydroxyl side-chain groups of D233 and S244, respectively (Fig. 3C). Last, Gal5 forms hydrogen bonds with both the S244 carbonyl backbone and the hydroxyl side-chain group. It also interacts with both the carboxyl group of the D233 side chain and the hydroxyl group of the S234 side chain through water-mediated hydrogen bonds (Fig. 3D). No interactions were observed between Gal2 and BabA.


Structural basis of Lewis(b) antigen binding by the Helicobacter pylori adhesin BabA.

Hage N, Howard T, Phillips C, Brassington C, Overman R, Debreczeni J, Gellert P, Stolnik S, Winkler GS, Falcone FH - Sci Adv (2015)

Interactions between BabA amino acids and Leb residues.(A) Fuc1 forms hydrogen bonds with the carbonyl backbone of C189, G191, and N194 and the side chain of T246. (B) Fuc4 forms a water-mediated hydrogen bond with the N206 side chain. (C) GlcNAc3 forms hydrogen bonds with the D233 and S244 side chains. (D) Gal5 forms hydrogen bonds with the carbonyl backbone and side chain of S244. Gal5 also forms hydrogen bonds with a water molecule structured by the side chains of D233 and S234. Fucose, galactose, and N-acetylglucosamine residues are colored orange, yellow, and blue, respectively. Hydrogen bonds are represented by dotted black lines.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4643811&req=5

Figure 3: Interactions between BabA amino acids and Leb residues.(A) Fuc1 forms hydrogen bonds with the carbonyl backbone of C189, G191, and N194 and the side chain of T246. (B) Fuc4 forms a water-mediated hydrogen bond with the N206 side chain. (C) GlcNAc3 forms hydrogen bonds with the D233 and S244 side chains. (D) Gal5 forms hydrogen bonds with the carbonyl backbone and side chain of S244. Gal5 also forms hydrogen bonds with a water molecule structured by the side chains of D233 and S234. Fucose, galactose, and N-acetylglucosamine residues are colored orange, yellow, and blue, respectively. Hydrogen bonds are represented by dotted black lines.
Mentions: The crystallographic model indicated that binding is mediated by a network of hydrogen bonds between Leb Fuc1, GlcNAc3, Fuc4, and Gal5 residues and a total of eight BabA amino acids. Fuc1 forms hydrogen bonds with the carbonyl backbone groups of C189, G191, and N194, and the hydroxyl group of the T246 side chain (Fig. 3A). Fuc4 interacts with the hydroxyl group of the N206 side chain through a water-mediated hydrogen bond (Fig. 3B). The GlcNAc3 residue forms two hydrogen bonds with the carboxyl and hydroxyl side-chain groups of D233 and S244, respectively (Fig. 3C). Last, Gal5 forms hydrogen bonds with both the S244 carbonyl backbone and the hydroxyl side-chain group. It also interacts with both the carboxyl group of the D233 side chain and the hydroxyl group of the S234 side chain through water-mediated hydrogen bonds (Fig. 3D). No interactions were observed between Gal2 and BabA.

Bottom Line: No conformational change occurs in BabA upon binding of Le(b), which is characterized by low affinity under acidic [K D (dissociation constant) of ~227 μM] and neutral (K D of ~252 μM) conditions.Binding is mediated by a network of hydrogen bonds between Le(b) Fuc1, GlcNAc3, Fuc4, and Gal5 residues and a total of eight BabA amino acids (C189, G191, N194, N206, D233, S234, S244, and T246) through both carbonyl backbone and side-chain interactions.The structural model was validated through the generation of two BabA variants containing N206A and combined D233A/S244A substitutions, which result in a reduction and complete loss of binding affinity to Le(b), respectively.

View Article: PubMed Central - PubMed

Affiliation: School of Pharmacy, University of Nottingham, University Park, Nottingham NG7 2RD, UK.

ABSTRACT
Helicobacter pylori is a leading cause of peptic ulceration and gastric cancer worldwide. To achieve colonization of the stomach, this Gram-negative bacterium adheres to Lewis(b) (Le(b)) antigens in the gastric mucosa using its outer membrane protein BabA. Structural information for BabA has been elusive, and thus, its molecular mechanism for recognizing Le(b) antigens remains unknown. We present the crystal structure of the extracellular domain of BabA, from H. pylori strain J99, in the absence and presence of Le(b) at 2.0- and 2.1-Å resolutions, respectively. BabA is a predominantly α-helical molecule with a markedly kinked tertiary structure containing a single, shallow Le(b) binding site at its tip within a β-strand motif. No conformational change occurs in BabA upon binding of Le(b), which is characterized by low affinity under acidic [K D (dissociation constant) of ~227 μM] and neutral (K D of ~252 μM) conditions. Binding is mediated by a network of hydrogen bonds between Le(b) Fuc1, GlcNAc3, Fuc4, and Gal5 residues and a total of eight BabA amino acids (C189, G191, N194, N206, D233, S234, S244, and T246) through both carbonyl backbone and side-chain interactions. The structural model was validated through the generation of two BabA variants containing N206A and combined D233A/S244A substitutions, which result in a reduction and complete loss of binding affinity to Le(b), respectively. Knowledge of the molecular basis of Le(b) recognition by BabA provides a platform for the development of therapeutics targeted at inhibiting H. pylori adherence to the gastric mucosa.

No MeSH data available.


Related in: MedlinePlus