Zawadzka Anna M, Abergel Rebecca J, Nichiporuk Rita, Andersen Ulla N, Raymond Kenneth N
Department of Chemistry, University of California, Berkeley, 94720-1460, USA.
Biochemistry. 2009 Apr 28;48(16):3645-57. doi: 10.1021/bi8018674.
During growth under iron limitation, Bacillus cereus and Bacillus anthracis, two human pathogens from the Bacillus cereus group of Gram-positive bacteria, secrete two siderophores, bacillibactin (BB) and petrobactin (PB), for iron acquisition via membrane-associated substrate-binding proteins (SBPs) and other ABC transporter components. Since PB is associated with virulence traits in B. anthracis, the PB-mediated iron uptake system presents a potential target for antimicrobial therapies; its characterization in B. cereus is described here. Separate transporters for BB, PB, and several xenosiderophores are suggested by (55)Fe-siderophore uptake studies. The PB precursor, 3,4-dihydroxybenzoic acid (3,4-DHB), and the photoproduct of FePB (FePB(nu)) also mediate iron delivery into iron-deprived cells. Putative SBPs were recombinantly expressed, and their ligand specificity and binding affinity were assessed using fluorescence spectroscopy. The noncovalent complexes of the SBPs with their respective siderophores were characterized using ESI-MS. The differences between solution phase behavior and gas phase measurements are indicative of noncovalent interactions between the siderophores and the binding sites of their respective SBPs. These studies combined with bioinformatics sequence comparison identify SBPs from five putative transporters specific for BB and enterobactin (FeuA), 3,4-DHB and PB (FatB), PB (FpuA), schizokinen (YfiY), and desferrioxamine and ferrichrome (YxeB). The two PB receptors show different substrate ranges: FatB has the highest affinity for ferric 3,4-DHB, iron-free PB, FePB, and FePB(nu), whereas FpuA is specific to only apo- and ferric PB. The biochemical characterization of these SBPs provides the first identification of the transporter candidates that most likely play a role in the B. cereus group pathogenicity.
在铁限制条件下生长时，蜡样芽孢杆菌和炭疽芽孢杆菌这两种革兰氏阳性菌蜡样芽孢杆菌属的人类病原体，会分泌两种铁载体，杆菌铁载体（BB）和石油铁载体（PB），通过膜相关底物结合蛋白（SBP）和其他ABC转运蛋白成分来获取铁。由于PB与炭疽芽孢杆菌的毒力特性相关，PB介导的铁摄取系统是抗菌治疗的一个潜在靶点；本文描述了其在蜡样芽孢杆菌中的特征。通过（55）Fe-铁载体摄取研究表明存在分别用于BB、PB和几种异源铁载体的转运蛋白。PB前体3,4-二羟基苯甲酸（3,4-DHB）以及FePB的光产物（FePB(nu)）也介导铁向缺铁细胞的传递。推定的SBP被重组表达，并使用荧光光谱法评估其配体特异性和结合亲和力。使用电喷雾电离质谱（ESI-MS）对SBP与其各自铁载体的非共价复合物进行了表征。溶液相行为和气相测量之间的差异表明铁载体与其各自SBP结合位点之间存在非共价相互作用。这些研究与生物信息学序列比较相结合，确定了来自五个推定转运蛋白的SBP，这些转运蛋白分别对BB和肠杆菌素（FeuA）、3,4-DHB和PB（FatB）、PB（FpuA）、裂殖菌素（YfiY）以及去铁胺和铁铬素（YxeB）具有特异性。两种PB受体显示出不同的底物范围：FatB对三价3,4-DHB、无铁PB、FePB和FePB(nu)具有最高亲和力，而FpuA仅对脱辅基PB和三价PB具有特异性。这些SBP的生化特性首次鉴定出了最有可能在蜡样芽孢杆菌属致病性中发挥作用的转运蛋白候选物。
