Metal binding spectrum and model structure of theBacillus anthracisvirulence determinant MntA

Abstract

The potentially lethal human pathogen Bacillus anthracis expresses a putative metal import system, MntBCA, which belongs to the large family of ABC transporters. MntBCA is essential for virulence of Bacillus anthracis: deletion of MntA, the system's substrate binding protein, yields a completely non-virulent strain. Here we determined the metal binding spectrum of MntA. In contrast to what can be inferred from growth complementation studies we find no evidence that MntA binds Fe(2+) or Fe(3+). Rather, MntA binds a variety of other metal ions, including Mn(2+), Zn(2+), Cd(2+), Co(2+), and Ni(2+) with affinities ranging from 10(-6) to 10(-8) M. Binding of Zn(2+) and Co(2+) have a pronounced thermo-stabilizing effect on MntA, with Mn(2+) having a milder effect. The thermodynamic stability of MntA, competition experiments, and metal binding and release experiments all suggest that Mn(2+) is the metal that is likely transported by MntBCA and is therefore the limiting factor for virulence of Bacillus anthracis. A homology-model of MntA shows a single, highly conserved metal binding site, with four residues that participate in metal coordination: two histidines, a glutamate, and an aspartate. The metals bind to this site in a mutually exclusive manner, yet surprisingly, mutational analysis shows that for proper coordination each metal requires a different subset of these four residues. ConSurf evolutionary analysis and structural comparison of MntA and its homologues suggest that substrate binding proteins (SBPs) of metal ions use a pair of highly conserved prolines to interact with their cognate ABC transporters. This proline pair is found exclusively in ABC import systems of metal ions.

Zn(II) suppresses biofilm formation in Bacillus amyloliquefaciens by inactivation of the Mn(II) uptake

“…At low intercellular concentrations of Mn(II), the mntABCD operons were assigned as the Mn(II) import system (Que and Helmann ). Our data indicate that despite Zn(II) and Mn(II) had similar binding affinities with MntA, Mn(II) is released much more readily than Zn(II), which is in accordance with a recent study, in which it was found that Zn(II) and Mn(II) bind differently to MntA in B. anthracis (Vigonsky et al ., ). Meanwhile, metal ion discrimination within the MntA is proposed to occur through subtle differences in the binding sites.…”

Substrate recognition and ATPase activity of the E. coli cysteine/cystine ABC transporter YecSC-FliY

“…We hypothesized that D-cysteine binds at the same site as Lcysteine or L-cystine, but that binding of Dcysteine induces a distinct conformational change that does not lead to increased thermostability. Recent studies have indeed demonstrated that binding of closely related substrates by SBPs can lead to different bound conformations (17,60,61). To explore this possibility, we conducted binding competition experiments using nanoDSF.…”

ATP binding and hydrolysis disrupt the high-affinity interaction between the heme ABC transporter HmuUV and its cognate substrate-binding protein

“…They are involved in many important physiological processes such as nutrient import, cellular detoxification, lipid homeostasis, signal transduction, antiviral defense, and antigen presentation (5)(6)(7)(8)(9)(10)(11). From a clinical perspective, ABC transporters are of great interest as they are directly involved in tumor resistance to multiple chemotherapeutics, bacterial multidrug resistance, and bacterial virulence and pathogenesis (12)(13)(14)(15)(16)(17)(18)(19). All ABC transporters share a basic architecture, minimally composed of two intracellular nucleotide-binding domains (NBDs), and two trans-membrane domains (TMDs).…”