Download MendeleyCrystal structures of an intrinsically active cholera toxin mutant yield insight into the toxin activation mechanism
O'Neal, C.J., Amaya, E.I., Jobling, M.G., Holmes, R.K., Hol, W.G.(2004) Biochemistry 43: 3772-3782
- PubMed: 15049684
- DOI: https://doi.org/10.1021/bi0360152
- Primary Citation of Related Structures:
1S5B, 1S5C, 1S5D, 1S5E, 1S5F - PubMed Abstract:
Cholera toxin (CT) is a heterohexameric bacterial protein toxin belonging to a larger family of A/B ADP-ribosylating toxins. Each of these toxins undergoes limited proteolysis and/or disulfide bond reduction to form the enzymatically active toxic fragment. Nicking and reduction render both CT and the closely related heat-labile enterotoxin from Escherichia coli (LT) unstable in solution, thus far preventing a full structural understanding of the conformational changes resulting from toxin activation. We present the first structural glimpse of an active CT in structures from three crystal forms of a single-site A-subunit CT variant, Y30S, which requires no activational modifications for full activity. We also redetermined the structure of the wild-type, proenzyme CT from two crystal forms, both of which exhibit (i) better geometry and (ii) a different A2 "tail" conformation than the previously determined structure [Zhang et al. (1995) J. Mol. Biol. 251, 563-573]. Differences between wild-type CT and active CTY30S are observed in A-subunit loop regions that had been previously implicated in activation by analysis of the structure of an LT A-subunit R7K variant [van den Akker et al. (1995) Biochemistry 34, 10996-11004]. The 25-36 activation loop is disordered in CTY30S, while the 47-56 active site loop displays varying degrees of order in the three CTY30S structures, suggesting that disorder in the activation loop predisposes the active site loop to a greater degree of flexibility than that found in unactivated wild-type CT. On the basis of these six new views of the CT holotoxin, we propose a model for how the activational modifications experienced by wild-type CT are communicated to the active site.
Organizational Affiliation:
Department of Chemistry and Biomolecular Structure Center, University of Washington, Seattle, Washington 98195, USA.
