Studies of a Ring-Cleaving Dioxygenase Illuminate the Role of Cholesterol Metabolism in the Pathogenesis of Mycobacterium tuberculosis

PLoS Pathogens

Paul J. Converse¹  Lindsay D. Eltis²  Haizhong Zhu²  Natalie Strynadka²  Katherine C. Yam²  Igor D'Angelo²  Jian-Xin Wang³  Victor Snieckus³  Lan H. Ly⁴  William R. Jacobs Jr.⁵  Rainer Kalscheuer⁵ 

[1] Center for Tuberculosis Research, Department of Medicine, Johns Hopkins University, Baltimore, Maryland, United States of America;Department of Biochemistry and Molecular Biology, University of British Columbia, Vancouver, British Columbia, Canada;Department of Chemistry, Queen's University, Kingston, Ontario, Canada;Department of Microbial and Molecular Pathogenesis, Texas A&M University Health Science Center, College Station, Texas, United States of America;Howard Hughes Medical Institute, Albert Einstein College of Medicine, Bronx, New York, United States of America

关键词: Cholesterol;    Mycobacterium tuberculosis;    Guinea pigs;    Enzyme metabolism;    Enzymes;    Crystal structure;    Electron density;    Metabolites;   

DOI  :  10.1371/journal.ppat.1000344

学科分类：生物科学（综合）

来源: Public Library of Science

【 摘 要 】

Mycobacterium tuberculosis, the etiological agent of TB, possesses a cholesterol catabolic pathway implicated in pathogenesis. This pathway includes an iron-dependent extradiol dioxygenase, HsaC, that cleaves catechols. Immuno-compromised mice infected with a ΔhsaC mutant of M. tuberculosis H37Rv survived 50% longer than mice infected with the wild-type strain. In guinea pigs, the mutant disseminated more slowly to the spleen, persisted less successfully in the lung, and caused little pathology. These data establish that, while cholesterol metabolism by M. tuberculosis appears to be most important during the chronic stage of infection, it begins much earlier and may contribute to the pathogen's dissemination within the host. Purified HsaC efficiently cleaved the catecholic cholesterol metabolite, DHSA (3,4-dihydroxy-9,10-seconandrost-1,3,5(10)-triene-9,17-dione; kcat/Km = 14.4±0.5 µM−1 s−1), and was inactivated by a halogenated substrate analogue (partition coefficient<50). Remarkably, cholesterol caused loss of viability in the ΔhsaC mutant, consistent with catechol toxicity. Structures of HsaC:DHSA binary complexes at 2.1 Å revealed two catechol-binding modes: bidentate binding to the active site iron, as has been reported in similar enzymes, and, unexpectedly, monodentate binding. The position of the bicyclo-alkanone moiety of DHSA was very similar in the two binding modes, suggesting that this interaction is a determinant in the initial substrate-binding event. These data provide insights into the binding of catechols by extradiol dioxygenases and facilitate inhibitor design.

【 授权许可】

CC BY