JB Accepts, published online ahead of print on 23 October 2009

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J. Bacteriol. doi:10.1128/JB.00930-09
Copyright (c) 2009, American Society for Microbiology and/or the Listed Authors/Institutions. All Rights Reserved.

Three-dimensional structure of different functional forms of Vibrio cholerae hemolysin oligomer: a cryo-electron microscopic study

Somnath Dutta, Budhaditya Mazumdar, Kalyan K. Banerjee, and Amar N. Ghosh^*

Division of Electron Microscopy, Division of Biochemistry, National Institute of Cholera and Enteric Diseases, P -33, C.I. T. Road, Scheme -XM, Beleghata, Kolkata-700010, India

^* To whom correspondence should be addressed. Email: ghoshan{at}icmr.org.in.

Vibrio cholerae hemolysin (HlyA) is a 65-kDa water-soluble pore-forming toxin that causes lysis of eukaryotic cells by destroying selective permeability of the plasma mambrane bilayer. The HlyA monomer self-assembles on the target cell surface to the more stable -barrel amphipathic heptamer which inserts into the membrane bilayer to form a diffusion channel. Deletion of the 15-kDa -prism lectin domain at the C-terminus generates a 50-kDa hemolysin variant (HlyA50) with 1000-fold decrease in hemolytic activity. Because functional differences are eventually dictated by structural differences, we determined three-dimensional structures of 65 and 50-kDa HlyA oligomers using cryo-electron microscopy and single particle methods. Our study clearly shows that the HlyA oligomer has 7-fold symmetry but HlyA50 oligomer is an asymmetric molecule. The HlyA oligomer has bowl-like, arm-like and ring-like domains. The bowl-like domain is coupled with the ring-like domain and seven sides opening are present just beneath the ring-like domain. Although a central channel is present in both HlyA and HlyA50 oligomers, they differ in pore-size as well as in shapes of the molecules and channel. These structural differences may be relevant to the striking difference in the efficiency of a functional channel formation by the two toxin forms.