Mechanistic insights into a novel exporter-importer system of Mycobacterium tuberculosis unravel its role in trafficking of iron.
BACKGROUND
Elucidation of the basic mechanistic and biochemical principles underlying siderophore mediated iron uptake in mycobacteria is crucial for targeting this principal survival strategy vis-à-vis virulence determinants of the pathogen. Although, an understanding of siderophore biosynthesis is known, the mechanism of their secretion and uptake still remains elusive.


RESULTS
Here, we demonstrate an interplay among three iron regulated Mycobacterium tuberculosis (M.tb) proteins, namely, Rv1348 (IrtA), Rv1349 (IrtB) and Rv2895c in export and import of M.tb siderophores across the membrane and the consequent iron uptake. IrtA, interestingly, has a fused N-terminal substrate binding domain (SBD), representing an atypical subset of ABC transporters, unlike IrtB that harbors only the permease and ATPase domain. SBD selectively binds to non-ferrated siderophores whereas Rv2895c exhibits relatively higher affinity towards ferrated siderophores. An interaction between the permease domain of IrtB and Rv2895c is evident from GST pull-down assay. In vitro liposome reconstitution experiments further demonstrate that IrtA is indeed a siderophore exporter and the two-component IrtB-Rv2895c system is an importer of ferrated siderophores. Knockout of msmeg_6554, the irtA homologue in Mycobacterium smegmatis, resulted in an impaired M.tb siderophore export that is restored upon complementation with M.tb irtA.


CONCLUSIONS
Our data suggest the interplay of three proteins, namely IrtA, IrtB and Rv2895c in synergizing the balance of siderophores and thus iron inside the mycobacterial cell.
Farhana
Aisha
A
Laboratory of Molecular and Cellular Biology, Centre for DNA Fingerprinting and Diagnostics, Hyderabad, India.
Kumar
Sandeep
S
Rathore
Shailendra S
SS
Ghosh
Prahlad C
PC
Ehtesham
Nasreen Z
NZ
Tyagi
Anil K
AK
Hasnain
Seyed E
SE
D018528:ATP-Binding Cassette Transporters / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D001426:Bacterial Proteins / Q000737:chemistry / Q000378:metabolism; D002462:Cell Membrane / Q000235:genetics / Q000378:metabolism; D016680:Genome, Bacterial; D007501:Iron / Q000378:metabolism*; D008264:Macrophages / Q000382:microbiology; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D009169:Mycobacterium tuberculosis / Q000235:genetics / Q000378:metabolism* / Q000472:pathogenicity; D009876:Operon; D016133:Polymerase Chain Reaction; D014774:Virulence
C437164:ABC transporter BcatrB; D018528:ATP-Binding Cassette Transporters; D001426:Bacterial Proteins; D008565:Membrane Proteins; D007501:Iron