In vitro characterization of a bacterial manganese uptake regulator of the Fur superfamily

Pierdomenico Bellini, Andrew M. Hemmings

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25 Citations (Scopus)


Fur proteins generally act as negative transcriptional regulators by binding to target regulatory sequences (fur boxes) in the promoter regions of iron-responsive genes. Recently, Rhizobium leguminosarum was reported to contain a protein (MurRl) of Fur-like sequence, which, under manganese-replete conditions in its native background, repressed transcription of an ABC-type Mn(II) transporter by binding to two nonpalindromic mur boxes in its promoter region. MurRl displays apparently unusual regulatory flexibility in that it can also repress iron-responsive genes in Escherichia coli under iron-replete conditions. In this study, we quantify the affinities for binding a number of first-row transition-metal cations by MurRl and demonstrate that, in a fashion similar to E. coli Fur, MurRl binds Mn(II), Fe(II), Zn(II), and Co(II) with similar micromolar-order dissociation constants. In contrast to the vast majority of Fur proteins, however, MurRl lacks any high-affinity structural Zn(II) sites. Furthermore, we show that holoMurRl binds as one and two homodimers to both mur and fur boxes in a concentration-dependent fashion in the presence of not only Mn(II) and Fe(II) but also Zn(II) and Co(II). We have developed an analytical method for determination of the individual dissociation constants and find that the DNA-binding affinities are essentially independent of the metal co-effector. These results complement those obtained in vivo by other authors and suggest that the Fur-like protein of R. leguminosarum, a competent ferric uptake regulator in E. coli, is insufficiently discriminating in its metal-binding characteristics to function as a regulator of iron homeostasis in its native background.
Original languageEnglish
Pages (from-to)2686-2698
Number of pages13
Issue number8
Publication statusPublished - 3 Feb 2006

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