1IKP

Pseudomonas Aeruginosa Exotoxin A, P201Q, W281A mutant

Summary for 1IKP

• Entry DOI: 10.2210/pdb1ikp/pdb
• Related: 1AER 1DMA 1ikq
• Descriptor: EXOTOXIN A, CHLORIDE ION, SODIUM ION, ... (4 entities in total)
• Functional Keywords: all 3 exotoxin a domains, transferase
• Biological source: Pseudomonas aeruginosa
• Total number of polymer chains: 1
• Total formula weight: 66904.71

Authors

McKay, D.B.,Wedekind, J.E.,Trame, C.B. (deposition date: 2001-05-04, release date: 2001-12-12, Last modification date: 2024-10-16)

Primary citation

Wedekind, J.E.,Trame, C.B.,Dorywalska, M.,Koehl, P.,Raschke, T.M.,McKee, M.,FitzGerald, D.,Collier, R.J.,McKay, D.B.
Refined Crystallographic Structure of Pseudomonas aeruginosa Exotoxin A and its Implications for the Molecular Mechanism of Toxicity
J.Mol.Biol., 314:823-837, 2001

PubMed Abstract: Exotoxin A of Pseudomonas aeruginosa asserts its cellular toxicity through ADP-ribosylation of translation elongation factor 2, predicated on binding to specific cell surface receptors and intracellular trafficking via a complex pathway that ultimately results in translocation of an enzymatic activity into the cytoplasm. In early work, the crystallographic structure of exotoxin A was determined to 3.0 A resolution, revealing a tertiary fold having three distinct structural domains; subsequent work has shown that the domains are individually responsible for the receptor binding (domain I), transmembrane targeting (domain II), and ADP-ribosyl transferase (domain III) activities, respectively. Here, we report the structures of wild-type and W281A mutant toxin proteins at pH 8.0, refined with data to 1.62 A and 1.45 A resolution, respectively. The refined models clarify several ionic interactions within structural domains I and II that may modulate an obligatory conformational change that is induced by low pH. Proteolytic cleavage by furin is also obligatory for toxicity; the W281A mutant protein is substantially more susceptible to cleavage than the wild-type toxin. The tertiary structures of the furin cleavage sites of the wild-type and W281 mutant toxins are similar; however, the mutant toxin has significantly higher B-factors around the cleavage site, suggesting that the greater susceptibility to furin cleavage is due to increased local disorder/flexibility at the site, rather than to differences in static tertiary structure. Comparison of the refined structures of full-length toxin, which lacks ADP-ribosyl transferase activity, to that of the enzymatic domain alone reveals a salt bridge between Arg467 of the catalytic domain and Glu348 of domain II that restrains the substrate binding cleft in a conformation that precludes NAD+ binding. The refined structures of exotoxin A provide precise models for the design and interpretation of further studies of the mechanism of intoxication.

PubMed: 11734000
DOI: 10.1006/jmbi.2001.5195

Experimental method

X-RAY DIFFRACTION (1.45 Å)