arsenic redux

What better way to spend a brisk december morning than by probing deeper into the landmark isolation of a bacteria that is able to grow in the absence of phosphorus?

The paper starts by noting the fundamental nutritional elemental requirements of all prokaryotic and eukaryotic cells: carbon, hydrogen, nitrogen, oxygen, sulfur, and phosphorus. (Other trace elements are necessary, often as enzyme co-factors, but are generally interchangeable) Arsenic lies below phosphorus on the periodic table giving it similar valency (3-) which contributes to its toxicity as it is mistakenly incorporated into metabolic pathways. Unfortunately, unlike chemically stable phosphorus-based interactions, arsenate-mediated bonds are much more easily hydrolyzed giving them a half-life too short to generate stable compounds. However, researchers the NASA Astrobiology Institute wondered whether, in certain nutrient-starved climates, bacteria may have adapted to substitute arsenic for phosphorus despite the inherent instability of arsenate compounds.

Their search began in Mono Lake (left), a water body with unusually high arsenic concentrations that would facilitate directed evolution of organisms that could cope with the inherent instability of arsenate-containing compounds. They inoculated media of pH 9.8 supplemented with glucose, vitamins, and trace metals but no phosphorus and increasing arsenate concentrations. They note that the inoculations underwent “many decimal dilution transfers greatly reducing any potential carryover of phosphorus.” The background phosphate in the medium was 3 micromolar, due to “trace impurities in the major salts.” They then isolated a growth which was reintroduced into phosphate-free medium, which grew out a strain, GFAJ-1, a member of the halomonadaceae family of gammaproteobacteria. The gammaproteobacteria are a large class of gram negative bacteria which include escherichia coli, psuedomonas aeruginosa, vibrio cholera, salmonella typherium, and yersinia species. The halomonadaceae family, specifically, has previously been shown to accumulate intracellular arsenic and has been suggested as a potential therapeutic agent for bioremediation of arsenic-contaminated water.

It is notable that this bacteria required either arsenic or phosphorus – it grew faster with phosphorus but was able to grow with exclusively arsenic. The bacteria grown in arsenic-exclusive media were larger in size (right, C vs. D) by scanning electron microscopy, potentially due to accumulation of large vacuoles seen on transmission EM (right, E). The authors appear to resolve the issue of phosphate impurity in the background media by showing that this bacteria could not grow in the As-/P- media, suggesting that the background phosphorus was insufficient to support growth in the absence of exogenous arsenic.

The group then went on to demonstrate arsenic incorporation into these bacteria; first by mass spectrometry of isolated bacteria and then by radiolabeled arsenic oxide, which appeared expectedly in protein, metabolite, lipid, and nucleic acid fractions (given this distribution of phosphate in other bacteria). The percentage of arsenic in nucleic acid, while small, appears to be consistent with the proportion of intracellular phosphate within nucleic acids – approximately 4% They then went on to show that arsenic was enriched in purified genomic DNA from bacteria grown in arsenic-exclusive media by ‘high-resolution secondary ion mass spectrometry’ (I don’t really know what this is). They then used “synchrotron X-ray studies” (what?) to assess how the arsenic was configured and found a structure consistent with As bound to 4 oxygen ions (similar to a phosphate group) as well as within small molecular weight metabolites (consistent with the role of phosphate in NADH, ATP, etc.) and modification of proteins on serine, threonine, and tyrosine residues (the main sites of protein phosphorylation – think of serine/threonine or tyrosine kinases).

Seems pretty solid to me.