Abstract
P1B-type ATPases are involved in heavy metal transport across the plasma membrane. Some Mycobacterium tuberculosis P-type ATPases are induced during infection, suggesting that this type of transporter could play a critical role in mycobacterial survival. To date, the ion specificity of M. tuberculosis heavy metal-transporting P1B-ATPases is not well understood. In this work, we observed that, although divalent heavy metal cations such as Cu2+, Co2+, Ni2+, Zn2+ Cd2+ and Pb2+ stimulate the ATPase activity of the putative P1B-type ATPase CtpG in the plasma membrane, whole cells of M. smegmatis expressing CtpG only tolerate high levels of Cd2+ and Cu2+. As indicator of the catalytic constant, Michaelis–Menten kinetics showed that CtpG embedded in the mycobacterial cell membrane has a V max/K m ratio 7.4-fold higher for Cd2+ than for Cu2+ ions. Thus, although CtpG can accept different substrates in vitro, this P-type ATPase transports Cd2+ more efficiently than other heavy metal cations across the mycobacterial plasma membrane.
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Acknowledgements
This work was supported by the División de Investigación Bogotá (DIB)-Universidad Nacional de Colombia (Grant 27754) and Colciencias (Grant 110171250419). ML and LQ were fellows of the “Jóvenes Investigadores e Innovadores” Program, Colciencias, Colombia.
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Communicated by Erko Stackebrandt.
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Supplementary Fig. 1 M. tuberculosis ctpG cloning in the pMV261 mycobacterial/E. coli shuttle vector. a Schematic representation of the pML01 recombinant expression vector. b The ctpG gene was amplified from M. tuberculosis H37Rv genomic DNA using the ctpG pMV-Dir/ ctpG pMV-Rev primers. Lanes: (1) Molecular weight marker (GeneRuler 1kb DNA Ladder, Thermo Scientific, USA) and (2) ctpG amplimer (2383 pb). c Integrity and directionality of ctpG in pML01 was verified by restriction patterns. Lanes: (1) Molecular weight marker (GeneRuler 1kb DNA Ladder, Thermo Scientific, USA) (2) pML01 recombinant vector, and restriction enzyme patterns obtained with the (3) NheI, (4) BglII, (5), XhoI, (6) EcoRI and (7) EcoRI/SalI restriction enzymes (DOCX 63 KB)
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López, M., Quitian, LV., Calderón, MN. et al. The P-type ATPase CtpG preferentially transports Cd2+ across the Mycobacterium tuberculosis plasma membrane. Arch Microbiol 200, 483–492 (2018). https://doi.org/10.1007/s00203-017-1465-z
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DOI: https://doi.org/10.1007/s00203-017-1465-z