Caractérisation des protéines ChuX, ChuY et ChuW de la voie d'acquisition de l'hème chez E. coli O157:H7

Authors: Labrie, Gabrielle
Advisor: Couture, Manon
Abstract: Pathogenic microorganisms that infect a mammalian host need iron to survive and grow. However, because of its low solubility at physiologic pH in aerobic environment, the majority of iron is sequestered in heme and proteins. Pathogenic bacteria have developed ways to acquire heme from the host as a source of iron. ChuX, ChuY and ChuW are proteins encoded by a gene cluster involved in direct heme acquisition in Escherichia coli O157:H7. So far, these three proteins have received little attention. In this study, the functions of these proteins have been investigated based on predictions from the primary sequence analyses. Our results reveal that the ChuY protein, belonging to the short chain dehydrogenase/reductase family, reduces the tripyrrole derived from the heme degradation reaction catalyzed by the ChuS protein using NADPH as cofactor. ChuY also accelerates nine times the rate-limiting step of the ChuS reaction, which is the opening of the porphyrin ring of verdoheme. Spectroscopic studies and the determination of the pre-stationary kinetic constants have confirmed the catalytic reductase activity of ChuY. From our results, a model of heme degradation involving both the ChuY and ChuS proteins is proposed. The ChuX protein is predicted to have a role in the transport and storage of heme. Our results show that holo-ChuX can transfer heme to the apo-ChuS protein quickly, after which heme degradation can start. Enzymatic testing of the ChuW protein could not be carried out because of the insolubility of this protein. Overall, this study has resulted in a better understanding of the respective role of the ChuY and ChuX proteins in the direct heme acquisition system of E. coli O157:H7.
Document Type: Mémoire de maîtrise
Issue Date: 2016
Open Access Date: 18 May 2018
Permalink: http://hdl.handle.net/20.500.11794/29751
Grantor: Université Laval
Collection:Thèses et mémoires

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