Wastewater effluents from mines and metal refineries are often contaminated with heavy metal ions, so they pose hazards to human and environmental health. Conventional technologies to remove heavy metal ions are well-established, but the most popular methods have drawbacks: chemical precipitation generates sludge waste, and activated carbon and ion exchange resins are made from unsustainable non-renewable resources. Using microbial biomass as the platform for heavy metal ion removal is an alternative method. Specifically, bioaccumulation is a natural biological phenomenon where microorganisms use proteins to uptake and sequester metal ions in the intracellular space to utilize in cellular processes (e.g., enzyme catalysis, signaling, stabilizing charges on biomolecules). Recombinant expression of these import-storage systems in genetically engineered microorganisms allows for enhanced uptake and sequestration of heavy metal ions. This has been studied for over two decades for bioremediative applications, but successful translation to industrial-scale processes is virtually non-existent. Meanwhile, demands for metal resources are increasing while discovery rates to supply primary grade ores are not. This review re-thinks how bioaccumulation can be used and proposes that it can be developed for bioextractive applications—the removal and recovery of heavy metal ions for downstream purification and refining, rather than disposal. This review consolidates previously tested import-storage systems into a biochemical framework and highlights efforts to overcome obstacles that limit industrial feasibility, thereby identifying gaps in knowledge and potential avenues of research in bioaccumulation.

Haloacid dehalogenase (HAD)-like hydrolases are a vast superfamily of largely uncharacterized enzymes, with a few members shown to possess phosphatase, β-phosphoglucomutase, phosphonatase, and dehalogenase activities. Using a representative set of 80 phosphorylated substrates, we characterized the substrate specificities of 23 soluble HADs encoded in the Escherichia coli genome. We identified small molecule phosphatase activity in 21 HADs and β-phosphoglucomutase activity in one protein. The E. coli HAD phosphatases show high catalytic efficiency and affinity to a wide range of phosphorylated metabolites that are intermediates of various metabolic reactions. Rather than following the classical “one enzyme-one substrate” model, most of the E. coli HADs show remarkably broad and overlapping substrate spectra. At least 12 reactions catalyzed by HADs currently have no EC numbers assigned in Enzyme Nomenclature. Surprisingly, most HADs hydrolyzed small phosphodonors (acetyl phosphate, carbamoyl phosphate, and phosphoramidate), which also serve as substrates for autophosphorylation of the receiver domains of the two-component signal transduction systems. The physiological relevance of the phosphatase activity with the preferred substrate was validated in vivo for one of the HADs, YniC. Many of the secondary activities of HADs might have no immediate physiological function but could comprise a reservoir for evolution of novel phosphatases.

Summary: Clustered Regularly Interspaced Short Palindromic Repeats (CRISPRs) and the associated proteins (Cas) comprise a system of adaptive immunity against viruses and plasmids in prokaryotes. Cas1 is a CRISPR‐associated protein that is common to all CRISPR‐containing prokaryotes but its function remains obscure. Here we show that the purified Cas1 protein of Escherichia coli (YgbT) exhibits nuclease activity against single‐stranded and branched DNAs including Holliday junctions, replication forks and 5′‐flaps. The crystal structure of YgbT and site‐directed mutagenesis have revealed the potential active site. Genome‐wide screens show that YgbT physically and genetically interacts with key components of DNA repair systems, including recB, recC and ruvB. Consistent with these findings, the ygbT deletion strain showed increased sensitivity to DNA damage and impaired chromosomal segregation. Similar phenotypes were observed in strains with deletion of CRISPR clusters, suggesting that the function of YgbT in repair involves interaction with the CRISPRs. These results show that YgbT belongs to a novel, structurally distinct family of nucleases acting on branched DNAs and suggest that, in addition to antiviral immunity, at least some components of the CRISPR–Cas system have a function in DNA repair.