A. Dias, D. Zamble
JBIC Journal of Biological Inorganic Chemistry, 2005
The Escherichia coli NikR is a 15-kDa protein that negatively regulates transcription of the nikABCDE operon that encodes for an ATP-dependent Ni(II) permease. Thermal and chemical denaturation studies with NikR previously demonstrated that Ni(II)–NikR is more stable than the protein bound to other metals such as Cu(II), Co(II) and Zn(II). To determine if Ni(II) induces a unique conformational change in NikR, digestion experiments with selected proteases were performed in the presence of the above metals. Both denaturing-polyacrylamide gel electrophoresis and reversed-phase HPLC revealed fragmentation patterns in the presence of stoichiometric nickel that were distinct from the cleavage of apo-NikR. Digestion of Cu(II)–NikR produced fragmentation that was similar, although less dramatic, to that produced with Ni(II)–NikR, whereas the Zn(II)- and Co(II)-bound proteins were digested in a similar manner as apo-NikR. Digestion fragments were collected, identified by MALDI-MS, and then mapped onto the available crystal structure of NikR. Although the specificity of the proteases utilized differed, the data suggest that Ni(II) has a selective allosteric effect and that upon metal binding the NikR metal-binding pocket is oriented or protected in such a way as to present itself for digestion in a unique conformation. This data sheds light on the Ni(II)-selective conformational changes that allow NikR to bind DNA optimally and repress transcription of the nik operon.
Cited by 6 publications.
Field of study: Chemistry