GFAJ-1 is a strain of rod-shaped bacteria in the family
Halomonadaceae . It is an extremophile that was isolated from the
hypersaline and alkaline
Mono Lake in eastern
Felisa Wolfe-Simon , a NASA research fellow in residence
US Geological Survey . In a 2010 Science journal publication,
the authors claimed that the microbe, when starved of phosphorus , is
capable of substituting arsenic for a small percentage of its
phosphorus to sustain its growth. Immediately after publication,
other microbiologists and biochemists expressed doubt about this claim
which was robustly criticized in the scientific community. Subsequent
independent studies published in 2012 found no detectable arsenate in
DNA of GFAJ-1, refuted the claim, and demonstrated that
simply an arsenate-resistant, phosphate-dependent organism.
* 1 Discovery
* 2 Taxonomy and phylogeny
Species or strain
* 3 Biochemistry
Arsenate ester stability
* 4 Criticism
* 5 See also
* 6 References
* 7 External links
Wolfe-Simon at Mono Lake, 2010
GFAJ-1 bacterium was discovered by geomicrobiologist Felisa
Wolfe-Simon , a NASA astrobiology fellow in residence at the US
Geological Survey in Menlo Park,
California . GFAJ stands for "Give
Felisa a Job". The organism was isolated and cultured beginning in
2009 from samples she and her colleagues collected from sediments at
the bottom of
Mono Lake , California, U.S.A.
Mono Lake is hypersaline
(about 90 grams/liter) and highly alkaline (pH 9.8). It also has one
of the highest natural concentrations of arsenic in the world (200 μM
). The discovery was widely publicized on 2 December 2010.
TAXONOMY AND PHYLOGENY
Escherichia coli strain O157:H7
Halomonas venusta strain NBSL13
Halomonas sp. GTW
Halomonas sp. G27
Halomonas sp. DH77
Halomonas sp. mp3
Halomonas sp. IB-O18
Halomonas sp. ML-185
GFAJ-1 based on ribosomal
Molecular analysis based on 16S r
RNA sequences shows
GFAJ-1 to be
closely related to other moderate halophile ("salt-loving") bacteria
of the family
Halomonadaceae . Although the authors produced a
cladogram in which the strain is nested among members of
including H. alkaliphila and H. venusta , they did not explicitly
assign the strain to that genus. Many bacteria are known to be able
to tolerate high levels of arsenic, and to have a proclivity to take
it up into their cells. However,
GFAJ-1 was controversially proposed
to go a step further; when starved of phosphorus, it was proposed to
instead incorporate arsenic into its metabolites and macromolecules
and continue growing.
The sequence of the genome of the bacterium
GFAJ-1 is now posted in
SPECIES OR STRAIN
Tufa formations along the shore of
In the Science journal article,
GFAJ-1 is referred to as a strain of
Halomonadaceae and not as a new species . The International Code of
Bacteria , the set of regulations which govern the
taxonomy of bacteria, and certain articles in the International
Journal of Systematic and Evolutionary
Microbiology contain the
guidelines and minimal standards to describe a new species, e.g. the
minimal standards to describe a member of the Halomonadaceae.
Organisms are described as new species if they meet certain
physiological and genetic conditions, such as generally less than 97%
RNA sequence identity to other known species and metabolic
differences allowing them to be discerned apart. In addition to
indicators to tell the novel species from other species, other
analyses are required, such as fatty acid composition , respiratory
quinone used and tolerance ranges and deposition of the strain in at
least two microbiological repositories. New proposed names are given
in italics followed by sp. nov. (and gen. nov. if it is a novel genus
according to the descriptions of that clade ). In the instance of
GFAJ-1 strain these criteria are not met, and the strain is not
claimed to be a new species. When a strain is not assigned to a
species (e.g. due to insufficient data or choice) it is often labeled
as the genus name followed by "sp." (i.e., undetermined species of
that genus) and the strain name. In the case of
GFAJ-1 the authors
chose to refer to the strain by strain designation only. Strains
closely related to
Halomonas sp. GTW and
G27, neither of which were described as valid species. If the
authors had formally assigned strain
GFAJ-1 to the
the name would be given as
Halomonas sp. GFAJ-1.
A phosphorus-free growth medium (which actually contained 3.1 ± 0.3
μM of residual phosphate, from impurities in reagents) was used to
culture the bacteria in a regime of increasing exposure to arsenate ;
the initial level of 0.1 mM was eventually ramped up to 40 mM.
Alternative media used for comparative experiments contained either
high levels of phosphate (1.5 mM) with no arsenate, or had neither
added phosphate nor added arsenate. It was observed that
grow through many doublings in cell numbers when cultured in either
phosphate or arsenate media, but could not grow when placed in a
medium of a similar composition to which neither phosphate nor
arsenate was added. The phosphorus content of the arsenic-fed,
phosphorus-starved bacteria (as measured by ICP-MS) was only 0.019 (±
0.001) % by dry weight, one thirtieth of that when grown in
phosphate-rich medium. This phosphorus content was also only about one
tenth of the cells' average arsenic content (0.19 ± 0.25% by dry
weight). The arsenic content of cells as measured by ICP-MS varies
widely and can be lower than the phosphorus contents in some
experiments, and up to fourteen times higher in others. Other data
from the same study obtained with nano-SIMS suggest a ~75-fold excess
of phosphate (P) over arsenic (As) when expressed as P:C and As:C
ratios, even in cells grown with arsenate and no added phosphate.
When cultured in the arsenate solution,
GFAJ-1 only grew 60% as fast
as it did in phosphate solution. The phosphate-starved bacteria had
an intracellular volume 1.5 times normal; the greater volume appeared
to be associated with the appearance of large "vacuole -like regions".
Scanning electron micrograph of
GFAJ-1 cells grown in defined
minimal medium supplemented with 1.5 mM phosphate
When the researchers added isotope-labeled arsenate to the solution
to track its distribution , they found that arsenic was present in the
cellular fractions containing the bacteria's proteins , lipids and
metabolites such as ATP , as well as its
RNA . Nucleic acids
from stationary phase cells starved of phosphorus were concentrated
via five extractions (one with phenol , three with phenol-chloroform
and one with chloroform extraction solvent), followed by ethanol
precipitation . Although direct evidence of the incorporation of
arsenic into biomolecules is still lacking, radioactivity measurements
suggested that approximately one-tenth (11.0 ± 0.1%) of the arsenic
absorbed by these bacteria ended up in the fraction that contained the
nucleic acids (
DNA and RNA) and all other co-precipitated compounds
not extracted by the previous treatments. A comparable control
experiment with isotope-labeled phosphate was not performed. With the
distribution of the strain in mid-2011, other labs began to
independently test the validity of the discovery. Rosemary Redfield
University of British Columbia , following issues with the
growth conditions, investigated the growth requirements of GFAJ-1, and
found that the strain grows better on solid agar medium than in liquid
culture. Redfield attributed this to low potassium levels and
hypothesized that the potassium levels in basal ML60 medium may be too
low to support growth. Redfield after finding and addressing further
issues (ionic strength, pH and the use of glass tubes instead of
polypropylene) found that arsenate marginally stimulated growth, but
didn't affect the final densities of the cultures, unlike what was
claimed. Subsequent studies using mass spectrometry by the same group
found no evidence of arsenate being incorporated into the
ARSENATE ESTER STABILITY
Structure of poly-β-hydroxybutyrate
Arsenate esters , such as those that would be present in
DNA , are
generally expected to be orders of magnitude less stable to hydrolysis
than corresponding phosphate esters . dAMAs, the structural arsenic
analog of the
DNA building block dAMP , has a half-life of 40 minutes
in water at neutral pH. Estimates of the half-life in water of
arsenodiester bonds, which would link the nucleotides together, are as
short as 0.06 seconds—compared to 30 million years for the
phosphodiester bonds in DNA. The authors speculate that the bacteria
may stabilize arsenate esters to a degree by using
poly-β-hydroxybutyrate (which has been found to be elevated in
"vacuole-like regions" of related species of the genus
Halomonas ) or
other means to lower the effective concentration of water.
Polyhydroxybutyrates are used by many bacteria for energy and carbon
storage under conditions when growth is limited by elements other than
carbon, and typically appear as large waxy granules closely resembling
the "vacuole-like regions" seen in
GFAJ-1 cells. The authors present
no mechanism by which insoluble polyhydroxybutyrate may lower the
effective concentration of water in the cytoplasm sufficiently to
stabilize arsenate esters. Although all halophiles must reduce the
water activity of their cytoplasm by some means to avoid desiccation,
the cytoplasm always remains an aqueous environment.
NASA's announcement of a news conference "that will impact the search
for evidence of extraterrestrial life" was criticized as
sensationalistic and misleading; an editorial in New Scientist
commented "although the discovery of alien life, if it ever happens,
would be one of the biggest stories imaginable, this was light-years
In addition, many experts who have evaluated the paper have concluded
that the reported studies do not provide enough evidence to support
the claims made by the authors. In an online article on Slate ,
Carl Zimmer discussed the skepticism of several
scientists: "I reached out to a dozen experts ... Almost unanimously,
they think the NASA scientists have failed to make their case".
Steven A. Benner has expressed doubts that arsenate has
replaced phosphate in the
DNA of this organism. He suggested that the
trace contaminants in the growth medium used by Wolfe-Simon in her
laboratory cultures are sufficient to supply the phosphorus needed for
the cells' DNA. He believes that it is more likely that arsenic is
being sequestered elsewhere in the cells. University of British
Rosemary Redfield said that the paper "doesn't
present any convincing evidence that arsenic has been incorporated
DNA or any other biological molecule", and suggests that the
experiments lacked the washing steps and controls necessary to
properly validate their conclusions. Harvard microbiologist Alex
Bradley said that arsenic-containing
DNA would be so unstable in water
it could not have survived the analysis procedure.
On 8 December 2010, Science published a response by Wolfe-Simon, in
which she stated that criticism of the research was expected. In
response, a "
Frequently Asked Questions " page to improve
understanding of the work was posted on 16 December 2010. The team
plans to deposit the
GFAJ-1 strain in the ATCC and DSMZ culture
collections to allow widespread distribution. In late May 2011, the
strain has also been made available upon request directly from the
laboratory of the authors. Science has made the article freely
available. The article was published in print six months after
acceptance in the 3 June 2011 issue of Science. The publication was
accompanied by eight technical comments addressing various concerns
regarding the article's experimental procedure and conclusion,
as well as a response by the authors to these concerns. The editor
Bruce Alberts has indicated that some issues remain and that
their resolution is likely to be a long process. A review by Rosen et
al., in the March 2011 issue of the journal
Bioessays discusses the
technical issues with the Science paper, provides alternative
explanations, and highlights known biochemistry of other arsenic
resistant and arsenic utilizing microbes.
On 27 May 2011, Wolfe-Simon and her team responded to the criticism
in a follow-up Science journal publication. Then on January 2012 a
group of researchers led by Rosie Redfield at the University of
British Columbia analyzed the
GFAJ-1 using liquid
chromatography–mass spectrometry and could not detect any arsenic,
which Redfield calls a "clear refutation" of the original paper's
findings. A simple explanation for the
GFAJ-1 growth in medium
supplied with arsenate instead of phosphate was provided by a team of
researchers at the
University of Miami
University of Miami in Florida. After labeling the
ribosomes of a laboratory strain of
Escherichia coli with radioactive
isotopes, they followed bacterial growth in medium containing arsenate
but no phosphate. They found that arsenate induces massive degradation
of ribosomes , thus providing sufficient phosphate for the slow growth
of arsenate tolerant bacteria. Similarly, they suggest,
grow by recycling phosphate from degraded ribosomes, rather than by
replacing it with arsenate. Following the publication of the articles
challenging the conclusions of the original Science article first
describing GFAJ-1, the website
Retraction Watch argued that the
original article should be retracted because of misrepresentation of
Hypothetical types of biochemistry
Hypothetical types of biochemistry
Nucleic acid analogues
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