Bioinformatic Mapping of Opine-Like Zincophore Biosynthesis in Bacteria

Abstract

Zinc is an essential nutrient in biological systems due to its structural or catalytic requirement in proteins involved in diverse cellular processes. To meet this cellular demand, microbes must acquire sufficient zinc from their environment. However, many environments have low zinc availability. One of the mechanisms used by bacteria to acquire zinc is through the production of small molecules known as zincophores. Similar to bacterial siderophores used for iron uptake, zincophores are synthesized by the bacterium and exported and then reimported as zincophore-zinc complexes. Thus far, only four zincophores have been described, including two from the human pathogens Staphylococcus aureus and Pseudomonas aeruginosa, in which they play a critical role in zinc acquisition during infection, and one in a soil bacterium. To determine what other microbes may produce zincophores, we used bioinformatic analyses to identify new zincophore biosynthetic gene clusters (BGCs) and predict the diversity of molecules synthesized. Genome neighborhood network analysis identified approximately 250 unique zincophore-producing species from actinobacteria, firmicutes, proteobacteria, and fusobacteria. This indicates that zincophores are produced by diverse bacteria that inhabit a broad range of ecological niches. Many of the BGCs likely produce characterized zincophores, based on similarity to the characterized systems. However, this analysis also identified numerous BGCs that, based on the colocalization of additional modifying enzymes and sequence divergence of the biosynthetic enzymes, are likely to produce unique zincophores. Collectively, these findings provide a comprehensive understanding of the zincophore biosynthetic landscape that will be invaluable for future research on these important small molecules. IMPORTANCE Bacteria must acquire essential nutrients, including zinc, from their environment. For bacterial pathogens, this necessitates overcoming the host metal-withholding response known as nutritional immunity. A novel type of zinc uptake mechanism that involves the bacterial production of a small zinc-scavenging molecule was recently described in the human pathogens Staphylococcus aureus, Pseudomonas aeruginosa, and Yersinia pestis, as well as the soil-associated bacterium Paenibacillus mucilaginosus. This suggests that zincophores may be important for zinc acquisition in diverse environments. In this study, we sought to identify other zincophore-producing bacteria using bioinformatics. We identified almost 250 unique zincophore-producing species, including human and animal pathogens, as well as isolates from soil, rhizosphere, plant, and marine habitats. Crucially, we observed diversity at the amino acid and gene organization levels, suggesting that many of these species are producing unique zincophores. Together, our findings highlight the importance of zincophores for a broad array of bacteria living in diverse environments.

Zinc import mediated by AdcABC is critical for colonization of the dental biofilm by Streptococcus mutans in an animal model

“…This system is comprised of small molecules with high zinc affinity (zincophores) that are synthesized in the cytoplasm, exported, and re‐internalized as a zincophore–zinc complex by a dedicated import system (Grim et al., 2020). Despite evidence that zincophore‐mediated uptake systems are produced in a wide array of bacteria, a zincophore biosynthetic gene cluster as well as zincophore export and import genes are absent in streptococcal genomes (Morey & Kehl‐Fie, 2020). Collectively, our phenotypic characterizations and bioinformatic analysis indicate that the AdcABC transporter is the only known high‐affinity zinc import system of S. mutans .…”

The AdcACB/AdcAII system is Essential for Zinc Homeostasis and an Important Contributor of Enterococcus faecalis Virulence

“…In most bacterial pathogens, Zn homeostasis under the severe Zn restricted conditions that can be encountered in host environments depends on the activity of surface-associated Zn uptake systems from the ATP-binding cassette (ABC) transporter family (reviewed in (20)(21)(22)). In addition, important human pathogens such as Pseudomonas aeruginosa and Staphylococcus aureus produce Zn-binding metallophores, also known as zincophores (23).…”

Conformation of the Solute-Binding Protein AdcAII Influences Zinc Uptake in Streptococcus pneumoniae

“…To overcome host-imposed Zn(II) restriction, pathogenic bacteria have evolved high-affinity acquisition systems to compete for metal ions at the host-pathogen interface ( Ammendola et al., 2007 ; Sabri et al., 2009 ; Bayle et al., 2011 ; Pederick et al., 2015 ). These include the ubiquitous Type II ATP-binding cassette (ABC) family importers ( Lewis et al., 2012 ), zinc-iron permease (ZIP) transporters ( Grass et al., 2005 ; Karlinsey et al., 2010 ), P-type ATPases ( Lewinson et al., 2009 ; Chien et al., 2013 ), and the recently identified zincophore scavenging systems ( Lhospice et al., 2017 ; Morey and Kehl-Fie, 2020 ). Bacterial ABC permeases employ extra-cytoplasmic solute-binding proteins (SBPs) to obtain cargo such as Zn(II) ions from the periplasm (Gram-negative bacteria) or the extracellular environment (Gram-positive bacteria) ( Lewis et al., 2012 ).…”