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Autooxidation Properties of Mycobacterium tuberculosis DevS and Impact of the Y171F Mutation on Gas Binding Kinetics
Author(s) -
Ioanoviciu Alexandra Smaranda,
Meharenna Yergalem,
Poulos Thomas,
Ortiz de Montellano Paul
Publication year - 2009
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.23.1_supplement.503.1
Subject(s) - devs , chemistry , carbon monoxide , mycobacterium tuberculosis , nitric oxide , biophysics , mutant , photochemistry , biochemistry , biology , gene , organic chemistry , catalysis , modeling and simulation , medicine , tuberculosis , systems engineering , pathology , engineering
In Mycobacterium tuberculosis , DevS is responsible for DevR activation in conjunction with DosT. Exposure to unfavorable stimuli is translated via the DevS/DevR two‐component system into the expression of genes of the "dormancy regulon." The DevS kinase activity is modulated by the binding of nitric oxide, carbon monoxide, and oxygen to the heme iron. We have cloned and characterized both wild‐type DevS and the Y171F mutant. Mutation of tyrosine 171 to a phenyl alanine was previously shown to impact kinase activity. In the present study we examine diatomic gas binding kinetics to wild‐type DevS and Y171F DevS using stopped‐flow combined with UV‐visible spectroscopy and laser flash photolysis. The Y171F mutation increases the on‐rates of carbon monoxide and oxygen, probably due to the requirement for displacement of a hydrogen bonded water molecule located in the distal heme cavity of the reduced wild‐type protein. In addition, we examine the stability of the wild‐type DevS oxy‐complex in the presence of ubiquitous metal cations. Oxy‐DevS is stable to autooxidation, suggesting that it functions as a gas and not redox sensor in vivo . This work was supported by the National Institutes of Health through grant AI074824.