Traces of metal are required for fundamental biochemical processes, such as photosynthesis and respiration. Cyanobacteria metal homeostasis acquires an important role because the photosynthetic machinery imposes a high demand for metals, making them a limiting factor for cyanobacteria, especially in the open oceans. On the other hand, in the last two centuries, the metal concentrations in marine environments and lake sediments have increased as a result of several industrial activities. In all cases, cells have to tightly regulate uptake to maintain their intracellular concentrations below toxic levels. Mechanisms to obtain metal under limiting conditions and to protect cells from an excess of metals are present in cyanobacteria. Understanding metal homeostasis in cyanobacteria and the proteins involved will help to evaluate the use of these microorganisms in metal bioremediation. Furthermore, it will also help to understand how metal availability impacts primary production in the oceans. In this review, we will focus on copper, nickel, cobalt and arsenic (a toxic metalloid) metabolism, which has been mainly analyzed in model cyanobacterium Synechocystis sp. PCC 6803.

A bacterial strain able to use cyanide as the sole nitrogen source under alkaline conditions has been isolated. The bacterium was classified as Pseudomonas pseudoalcaligenes by comparison of its 16S RNA gene sequence to those of existing strains and deposited in the Colección Española de Cultivos Tipo (Spanish Type Culture Collection) as strain CECT5344. Cyanide consumption is an assimilative process, since (i) bacterial growth was concomitant and proportional to cyanide degradation and (ii) the bacterium stoichiometrically converted cyanide into ammonium in the presence of l -methionine- d , l -sulfoximine, a glutamine synthetase inhibitor. The bacterium was able to grow in alkaline media, up to an initial pH of 11.5, and tolerated free cyanide in concentrations of up to 30 mM, which makes it a good candidate for the biological treatment of cyanide-contaminated residues. Both acetate and d , l -malate were suitable carbon sources for cyanotrophic growth, but no growth was detected in media with cyanide as the sole carbon source. In addition to cyanide, P. pseudoalcaligenes CECT5344 used other nitrogen sources, namely ammonium, nitrate, cyanate, cyanoacetamide, nitroferricyanide (nitroprusside), and a variety of cyanide-metal complexes. Cyanide and ammonium were assimilated simultaneously, whereas cyanide strongly inhibited nitrate and nitrite assimilation. Cyanase activity was induced during growth with cyanide or cyanate, but not with ammonium or nitrate as the nitrogen source. This result suggests that cyanate could be an intermediate in the cyanide degradation pathway, but alternative routes cannot be excluded.

Cyanase catalyzes the decomposition of cyanate into CO ₂ and ammonium, with carbamate as an unstable intermediate. The cyanase of Pseudomonas pseudoalcaligenes CECT5344 was negatively regulated by ammonium and positively regulated by cyanate, cyanide, and some cyanometallic complexes. Cyanase activity was not detected in cell extracts from cells grown with ammonium, even in the presence of cyanate. Nevertheless, a low level of cyanase activity was detected in nitrogen-starved cells. The cyn gene cluster of P. pseudoalcaligenes CECT5344 was cloned and analyzed. The cynA, cynB , and cynD genes encode an ABC-type transporter, the cynS gene codes for the cyanase, and the cynF gene encodes a novel σ ⁵⁴ -dependent transcriptional regulator which is not present in other bacterial cyn gene clusters. The CynS protein was expressed in Escherichia coli and purified by following a simple and rapid protocol. The P. pseudoalcaligenes cyanase showed an optimal pH of 8.5°C and a temperature of 65°C. An insertion mutation was generated in the cynS gene. The resulting mutant was unable to use cyanate as the sole nitrogen source but showed the same resistance to cyanate as the wild-type strain. These results, in conjunction with the induction pattern of the enzymatic activity, suggest that the enzyme has an assimilatory function. Although the induction of cyanase activity in cyanide-degrading cells suggests that some cyanate may be generated from cyanide, the cynS mutant was not affected in its ability to degrade cyanide, which unambiguously indicates that cyanate is not a central metabolite in cyanide assimilation.

Cyanide is one of the most potent and toxic chemicals produced by industry. The jewellery industry of Córdoba (Spain) generates a wastewater (residue) that contains free cyanide, as well as large amounts of cyano–metal complexes. Cyanide is highly toxic to living systems because it forms very stable complexes with transition metals that are essential for protein function. In spite of its extreme toxicity, some organisms have acquired mechanisms to avoid cyanide poisoning. The biological assimilation of cyanide needs the concurrence of three separate processes: (i) a cyanide-insensitive respiratory chain, (ii) a system for iron acquisition (siderophores) and (iii) a cyanide assimilation pathway. Siderophores are low-molecular-mass compounds (600–1500 Da) that scavenge iron (Fe3+) ions (usually with extremely high affinity) from the environment under iron-limiting conditions. There are two main classes of siderophores: catechol and hydroxamate types. The catechol-type siderophores chelate ferric ion via a hydroxy group, whereas the hydroxamate-type siderophores bind iron via a carbonyl group with the adjacent nitrogen. In the presence of cyanide, bacterial proliferation requires this specific metal uptake system because siderophores are able to break down cyano–metal complexes. Pseudomonas pseudoalcaligenes CECT5344 is able to use free cyanide or cyano–metal complexes as nitrogen source. A proteomic approach was used for the isolation and identification, in this strain, of a protein that was induced in the presence of cyanide, namely CN0, that is involved in siderophore biosynthesis in response to cyanide. An overview of bacterial cyanide degradation pathways and the involvement of siderophores in this process are presented.

Summary: P seudomonas pseudoalcaligenes CECT 5344 is a Gram‐negative bacterium able to tolerate cyanide and to use it as the sole nitrogen source. We report here the first draft of the whole genome sequence of a P . pseudoalcaligenes strain that assimilates cyanide. Three aspects are specially emphasized in this manuscript. First, some generalities of the genome are shown and discussed in the context of other P seudomonadaceae genomes, including genome size, G + C content, core genome and singletons among other features. Second, the genome is analysed in the context of cyanide metabolism, describing genes probably involved in cyanide assimilation, like those encoding nitrilases, and genes related to cyanide resistance, like the cio genes encoding the cyanide insensitive oxidases. Finally, the presence of genes probably involved in other processes with a great biotechnological potential like production of bioplastics and biodegradation of pollutants also is discussed.