Structural basis for the recognition of muramyltripeptide by Helicobacter pylori Csd4, a D,L-carboxypeptidase
Hyoun Sook Kim1, Jieun Kim2, Ha Na Im2
1Research Institute of Pharmaceutical Sciences, College of Pharmacy, Seoul National University, Seoul 151-742, Republic of Korea.
Abstract:
Helicobacter pylori infection causes a variety of gastrointestinal diseases, including peptic ulcers and gastric cancer. Its colonization of the gastric mucosa of the human stomach is a prerequisite for survival in the stomach. Colonization depends on its motility, which is facilitated by the helical shape of the bacterium. In H. pylori, cross-linking relaxation or trimming of peptidoglycan muropeptides affects the helical cell shape. Csd4 has been identified as one of the cell shape-determining peptidoglycan hydrolases in H. pylori. It is a Zn(2+)-dependent D,L-carboxypeptidase that cleaves the bond between the γ-D-Glu and the mDAP of the non-cross-linked muramyltripeptide (muramyl-L-Ala-γ-D-Glu-mDAP) of the peptidoglycan to produce the muramyldipeptide (muramyl-L-Ala-γ-D-Glu) and mDAP. Here, the crystal structure of H. pylori Csd4 (HP1075 in strain 26695) is reported in three different states: the ligand-unbound form, the substrate-bound form and the product-bound form. H. pylori Csd4 consists of three domains: an N-terminal D,L-carboxypeptidase domain with a typical carboxypeptidase fold, a central β-barrel domain with a novel fold and a C-terminal immunoglobulin-like domain. The D,L-carboxypeptidase domain recognizes the substrate by interacting primarily with the terminal mDAP moiety of the muramyltripeptide. It undergoes a significant structural change upon binding either mDAP or the mDAP-containing muramyltripeptide. It it also shown that Csd5, another cell-shape determinant in H. pylori, is capable of interacting not only with H. pylori Csd4 but also with the dipeptide product of the reaction catalyzed by Csd4.
Insights
Helicobacter pylori Csd4, a peptidoglycan hydrolase, maintains bacterial shape for stomach colonization. Its crystal structure reveals how it binds substrates and interacts with other cell shape determinants like Csd5.
Area of Science:
- Microbiology
- Structural Biology
- Biochemistry
Background:
- Helicobacter pylori colonization of the gastric mucosa is crucial for causing gastrointestinal diseases.
- Bacterial motility, essential for colonization, depends on the helical shape of H. pylori.
- Peptidoglycan muropeptide processing by enzymes like Csd4 influences bacterial cell shape.
Purpose of the Study:
- To determine the crystal structure of H. pylori Csd4 in different functional states.
- To elucidate the substrate-binding mechanism of Csd4.
- To investigate the interaction between Csd4 and other cell shape determinants, such as Csd5.
Main Methods:
- X-ray crystallography to determine the 3D structure of H. pylori Csd4.
- Biochemical assays to analyze enzyme activity and substrate interactions.
- Analysis of protein-protein interactions between Csd4 and Csd5.
Main Results:
- The crystal structure of H. pylori Csd4 was determined in ligand-unbound, substrate-bound, and product-bound forms.
- Csd4 comprises three domains: D,L-carboxypeptidase, novel β-barrel, and immunoglobulin-like.
- The D,L-carboxypeptidase domain undergoes conformational changes upon substrate binding, primarily interacting with the mDAP moiety.
Conclusions:
- Csd4's structure provides insights into its enzymatic mechanism and role in maintaining H. pylori's helical shape.
- Csd4's interaction with its substrate and product is structurally characterized.
- Csd4 interacts with Csd5, suggesting a coordinated mechanism for cell shape determination in H. pylori.
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