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Infection and Immunity, August 2009, p. 3389-3401, Vol. 77, No. 8
0019-9567/09/$08.00+0 doi:10.1128/IAI.00143-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.
Mycobacterium tuberculosis Cpn60.2 and DnaK Are Located on the Bacterial Surface, Where Cpn60.2 Facilitates Efficient Bacterial Association with Macrophages
Tyler B. M. Hickey,1
Lisa M. Thorson,2
David P. Speert,1,2,3
Mamadou Daffé,4 and
Richard W. Stokes1,2,3*
Departments of Pathology and Laboratory Medicine,1 Paediatrics, University of British Columbia, Vancouver, British Columbia, Canada,2 Division of Infectious and Immunological Diseases, British Columbia's Children's Hospital, Vancouver, British Columbia, Canada,3 Department of Molecular Mechanisms of Mycobacterial Infections, Institut de Pharmacologie et de Biologie Structurale du Centre National de la Recherche Scientifique et de l'Université Paul Sabatier (UMR 5089), Toulouse, France4
Received 5 February 2009/ Returned for modification 6 March 2009/ Accepted 15 May 2009
Mycobacterium tuberculosis, the causative agent of tuberculosis, initially contacts host cells with elements of its outer cell wall, or capsule. We have shown that capsular material from the surface of M. tuberculosis competitively inhibits the nonopsonic binding of whole M. tuberculosis bacilli to macrophages in a dose-dependent manner that is not acting through a global inhibition of macrophage binding. We have further demonstrated that isolated M. tuberculosis capsular proteins mediate a major part of this inhibition. Two-dimensional polyacrylamide gel electrophoresis analysis of the capsular proteins showed the presence of a wide variety of protein species, including proportionately high levels of the Cpn60.2 (Hsp65, GroEL2) and DnaK (Hsp70) molecular chaperones. Both of these proteins were subsequently detected on the bacterial surface. To determine whether these molecular chaperones play a role in bacterial binding, recombinant Cpn60.2 and DnaK were tested for their ability to inhibit the association of M. tuberculosis bacilli with macrophages. We found that recombinant Cpn60.2 can inhibit 
57% of bacterial association with macrophages, while DnaK was not inhibitory at comparable concentrations. Additionally, when polyclonal F(ab')2 fragments of anti-Cpn60.2 and anti-DnaK were used to mask the surface presentation of these molecular chaperones, a binding reduction of 34% was seen for anti-Cpn60.2 F(ab')2, while anti-DnaK F(ab')2 did not significantly reduce bacterial association with macrophages. Thus, our findings suggest that while M. tuberculosis displays both surface-associated Cpn60.2 and DnaK, only Cpn60.2 demonstrates adhesin functionality with regard to macrophage interaction.
* Corresponding author. Mailing address: Child and Family Research Institute, 950 West 28th Avenue, Room 303, Vancouver, British Columbia, V5Z 4H4 Canada. Phone: (604) 875-2491. Fax: (604) 875-2226. E-mail: rstokes{at}interchange.ubc.ca
Published ahead of print on 26 May 2009.
Infection and Immunity, August 2009, p. 3389-3401, Vol. 77, No. 8
0019-9567/09/$08.00+0 doi:10.1128/IAI.00143-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.
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