Crystal structure confirmation of JHP933 as a nucleotidyltransferase superfamily protein from Helicobacter pylori strain J99

Abstract

Helicobacter pylori is a well-known pathogen involved in the development of peptic ulcer, gastric adenocarcinoma and other forms of gastric cancer. Recently, there has been more considerable interest in strain-specific genes located in plasticity regions with great genetic variability. However, little is known about many of these genes. Studies suggested that certain genes in this region may play key roles in the pathogenesis of H. pylori-associated gastroduodenal diseases. JHP933, a conserved putative protein of unknown function, is encoded by the gene in plasticity region of H. pylori strain J99. Here we have determined the structure of JHP933. Our work demonstrates that JHP933 is a nucleotidyltransferase superfamily protein with a characteristic αβαβαβα topology. A superposition demonstrates overall structural homology of the JHP933 N-terminal fragment with lincosamide antibiotic adenylyltransferase LinA and identifies a possible substrate-binding cleft of JHP933. Furthermore, through structural comparison with LinA and LinB, we pinpoint conservative active site residues which may contribute to divalent ion coordination and substrate binding.

Figure 1. Overall structure of JHP933.

Ribbon diagram of the JHP933 structure, N-terminal core domain is colored in lime and C-terminal tail domain in cyan. α-helices are labelled with α, β-strands are labelled with β, and 3₁₀ helices are labelled with η.

Figure 2. The superposition of JHP933 and LinA/Lincomycin complex (4E8J) structures.

Ribbon diagram of JHP933/LinA, with JHP933 is colored in lime and LinA in magenta, and substrate lincomycin of LinA is shown in ball-and-stick representation.

Figure 3. Sequence and secondary structure comparison of JHP933 with structurally related LinA.

The secondary structures of JHP93 (top row) are labeled in lime and LinA from S. haemolyticu (bottom row) in magenta. The conserved active site motifs involved in catalysis ([DE]h[DE]h, h[DE]h) and substrate binding (hG) of NTase superfamily are shadowed in gray.

Figure 4. Putative substrate binding site of JHP933.

Ribbon diagram and surface representation of JHP933 are colored in lime, the modelled substrate lincomycin of the superimposed LinA/lincomycin complex is shown in ball-and-stick representation and colored in magenta (LinA protein not shown).

Figure 5. Active site conservation and substrate binding of JHP933, LinA and LinB.

The C atoms of active site residues are shown in ball-and-stick representation and distinctively colored: lime for JHP933, magenta for LinA (4E8J), and cyan for LinB (3JZ0). The substrate Mg²⁺ ions, as cyan spheres, AMPCPP and clindamycin, in yellow, are from LinB complex structure.