AdcA and AdcAII employ distinct zinc acquisition mechanisms and contribute additively to zinc homeostasis in Streptococcus pneumoniae

Abstract

Streptococcus pneumoniae is a globally significant human pathogen responsible for nearly 1 million deaths annually. Central to the ability of S. pneumoniae to colonize and mediate disease in humans is the acquisition of zinc from the host environment. Zinc uptake in S. pneumoniae occurs via the ATP-binding cassette transporter AdcCB, and, unusually, two zinc-binding proteins, AdcA and AdcAII. Studies have suggested that these two proteins are functionally redundant, although AdcA has remained uncharacterized by biochemical methods. Here we show that AdcA is a zinc-specific substrate-binding protein (SBP). By contrast with other zinc-binding SBPs, AdcA has two zinc-binding domains: a canonical amino-terminal cluster A-I zinc-binding domain and a carboxy-terminal zinc-binding domain, which has homology to the zinc-chaperone ZinT from Gram-negative organisms. Intriguingly, this latter feature is absent from AdcAII and suggests that the two zinc-binding SBPs of S. pneumoniae employ different modalities in zinc recruitment. We further show that AdcAII is reliant upon the polyhistidine triad proteins for zinc in vitro and in vivo. Collectively, our studies suggest that, despite the overlapping roles of the two SBPs in zinc acquisition, they may have unique mechanisms in zinc homeostasis and act in a complementary manner during host colonization.

The Adc/Lmb System Mediates Zinc Acquisition in Streptococcus agalactiae and Contributes to Bacterial Growth and Survival

“…The particularity of the S. agalactiae Adc/Lmb transporter is the presence of an additional binding protein, Lmb, which is almost specific to human isolates. In S. agalactiae, Lmb, AdcA, and AdcAII are redundant for zinc acquisition in chemically defined medium, which is consistent with what was observed in S. pneumoniae (40). In human biological fluids, bacterial growth was not affected by the loss of the Lmb protein alone, whereas the ⌬lmb ⌬adcA ⌬adcAII mutant strain was clearly outcompeted by the wild-type strain in cerebrospinal and amniotic fluids.…”

“…2). The histidine-rich loop and ZinT domain of the S. pneumoniae AdcA homolog have been suggested to aid in recruiting Zn 2ϩ (40). The structural differences between S. agalactiae AdcA and Lmb/ AdcAII proteins are similar to those observed between S. pneumoniae AdcA and AdcAII proteins and correspond to distinct zinc acquisition mechanisms (40,41).…”

“…The AdcA/Lmb family proteins are conserved among most streptococci and share a high degree of homology with zinc-binding proteins, in particular the AdcA and AdcAII proteins of S. pneumoniae (Table 2), whose role in zinc transport has been extensively studied (16,39,40). Following sequence alignment, we observed that the residues forming the Zn 2ϩ ion-containing binding site of Lmb (His66, His142, His206, and Glu281) are conserved in the AdcA and AdcAII proteins (18) (Fig.…”

“…In the three pathogenic streptococci, S. pneumoniae, S. pyogenes, and S. gordonii, the Adc system is composed of one AdcCB translocon and two zinc-binding proteins, the genes of which are found either within an adcRCBA operon or independently of the adc cluster (21). In S. pneumoniae, either AdcA or AdcAII is sufficient for zinc acquisition during growth in vitro and for systemic virulence in vivo, but both are necessary for optimal nasopharynx colonization (40). A recent study, again in S. pneumoniae, showed that the absence of AdcAII but not AdcA negatively affected early colonization of the nasopharynx (55).…”

“…These genes encode polyhistidine triad proteins, and S. agalactiae Sht has been shown to promote complement degradation by binding to factor H (12). In S. pneumoniae, the Sht homologs, called phtproteins, aid in zinc delivery to the ABC transporter substrate-binding protein AdcAII (40,41) and play a role in pneumococcal adhesion to the respiratory epithelium (57). The involvement of S. agalactiae Sht proteins in zinc transport, their interaction with the Adc/Lmb system components, and their putative redundancy are now under study and should provide new insights to better understand mechanisms of zinc acquisition in S. agalactiae.…”