PMID:18662316

Citation	Kikkawa, HS, Ueda, T, Suzuki, S and Yasuda, J (2008) Characterization of the catalytic activity of the gamma-phage lysin, PlyG, specific for Bacillus anthracis. FEMS Microbiol. Lett. 286:236-40
Abstract	Bacillus anthracis causes anthrax, a lethal disease affecting humans that has attracted attention due to its bioterrorism potential. PlyG is a lysin of gamma-phage, which specifically infects B. anthracis and lyses its cell wall. PlyG contains a T7 lysozyme-like amidase domain, which appears to be the catalytic domain, in the N-terminal region and has a high degree of sequence similarity with PlyL, which is an N-acetylmuramoyl-l-alanine amidase encoded by the B. anthracis genome. Here, we demonstrated that two amino acid residues of PlyG, H29 and E90, are necessary for its catalytic activity in B. anthracis. These residues are structurally analogous to residues whose mutation in T7 lysozyme abolished its catalytic activity. A C-terminal deletion mutant of PlyG lacking the core sequence for binding to B. anthracis showed completely abolished binding activity, unlike PlyL, despite high sequence similarity with PlyL in the N-terminal region. This suggests that the C-terminal binding domain, as well as the N-terminal catalytic domain, is essential for the catalytic activity of PlyG. Our observations provide new insights into the mechanism of specific catalysis of PlyG in B. anthracis and may contribute to the establishment of new methods for anthrax therapy.
Links	PubMed Online version:10.1111/j.1574-6968.2008.01280.x
Keywords	Amidohydrolases/genetics; Amidohydrolases/metabolism; Amino Acid Sequence; Amino Acid Substitution; Bacillus Phages/enzymology; Bacillus anthracis/virology; Bacterial Proteins/genetics; Catalytic Domain; Molecular Sequence Data; Mutagenesis, Site-Directed; N-Acetylmuramoyl-L-alanine Amidase/genetics; N-Acetylmuramoyl-L-alanine Amidase/metabolism; Protein Binding; Protein Structure, Tertiary; Sequence Alignment; Sequence Deletion; Viral Proteins/genetics; Viral Proteins/metabolism
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Annotations

Gene product	Qualifier	GO Term	Evidence Code	with/from	Aspect	Extension	Notes	Status
9CAUD:Q8LTE6	Contributes to	GO:0009253: peptidoglycan catabolic process	ECO:0000247: sequence alignment evidence used in manual assertion	UniProtKB:BACAN	P		Figure 1a in the literature shows a schematic representation of PlyG and PlyL, and the umbers of amino acid residues involved in catalysis are respectively, 233 and 234, indicating a high sequence similarity and low E-value, allowing assumption of function similarity to the endolysin PlyL. In addition, figure 2a, shows a sequence alignment of PlyG and PlyL and PlyG has a high degree of sequence similarity to PlyL. There are only 10 amino acid residue differences within the N-terminal regions of PlyG and PlyL, while 30 amino acid residues are different within their C-terminal regions. Therefore, it seems likely that the N-terminal region of PlyG also has similar characteristics.	complete CACAO 12085
9CAUD:Q8LTE6		GO:0051672: catabolism by organism of cell wall peptidoglycan in other organism	ECO:0000315: mutant phenotype evidence used in manual assertion		P		Figure 2d. in the literature compares Lytic activities of wild-type PlyG and its C-terminal deletion mutant. Catalytic activity was examined by colony-forming assay, and the control, PlyGB, was set to 100%. As shown in the figure, deletion of the C-terminal region completely abolished the lytic ability of PlyG, with the colony forming units of the C-terminal deletion mutant at a very high percentage, versus the normal wild type PlyG, which yielded a very low colony forming unit percentage. This indicates that the cell-wall-binding domain of PlyG is essential for its catalytic activity, and that addition of PlyG restricts colonies of bacteria from forming, characterizing it as an endolysin with peptidoglycan catabolic process and cleavage.	complete CACAO 12086

References

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