From MicrobeWiki, the student-edited microbiology resource
A Microbial Biorealm page on the genus Haloarcula Marismortui
Higher order taxa
Cellular organisms; Archaea; Euryarchaeota; Halobacteria; Halobacteriales; Halobacteriaceae; Haloarcula
Description and significance
Haloarcula marismortui is a halophilic red Archaeon (from the Halobacteriaceae family) found in the Dead Sea, a high saline, low oxygen solubility, and high light intensity environment. Like other halophilic archaeal organisms, H. marismortui thrives in this extreme environment due to several adaptations in protein structure, metabolic strategies and physiologic responses. (2, 3)
It is important to have its genome sequenced because of its unique ability to survive in such an extreme environment. The three fold extremities listed above are not expected characteristics of an environment in which a particular organism is able to survive. Therefore, it is important to understand what physiologic responses are unique to the organism that allows it to thrive in such an extreme environment. This will allow researchers to understand the systems level mechanisms that underlie environmental response systems. Analyzing its genome also provides support for the previously proposed characteristics of halophilic archaea, like acidic proteome, as well as the evolution of their genome architecture. A better understanding of gene regulatory networks that influence protein-protein and protein-DNA interactions may also provide a framework for biotechnological applications in the future. See Application section below for a more detailed description. (2, 3)
It was isolated in the 1960s by Ginzburg et al. in the Dead Sea. It is very closely related to Haloarcula vallismortis, but differs in its cell morphology and ability to use different sugars and other compounds for function. (4)
H. marismortui has a genome that is 4275kb in size composed of nine replicons and 4242 protein coding genes. It is divided into high and low G+C content replicons. The large chromosome I is a 3132-kb replicon with a 62.36% G+C content. The other eight replicons are smaller, ranging from 33 to 410 kb with G+C contents ranging from 54.25% to 60.02%, averaging about 57%. This bipartite genome-content organization is generally found in all members of this group of organisms. The significance of this type of organization is unknown.
However, there are three small replicons in H. marismortui that encode functions that are essential for survival. Replicon pNG600 codes several genes that are found nowhere else in the genome of H. marismortuis: aconitase, a significant enzyme in the TCA cycle, a DNA polymerase B family protein, the large and small subunits of endonuclease VII, and two transcription factor B (TFB) orthologs. This replicon is also responsible for H. marismortuis’s ability to handle heavy metal stress. It encodes genes for about a dozen cation transport proteins that have various specificities, metal-ion dependent transcription regulators, and a mercuric reductase.
Replicon pNG700 codes for four essential enzymes used in folate metabolism: viz. methylenetetrahydrofolate dehydrogenase, 5, 10-methylentretrahydrofolate reductase, formyltetrahydrofolate synthetase and formimidoyltetrahydrofolate cyclodeaminase. It also codes for functions that act downstream to arginine breakdown.
Chromosome II is the third replicon that encodes essential functions. It encodes one of three rRNA operons, as well as carbamoyl phosphate synthase, succinate-semialdehyde dehydrogenase, pyruvate dehydrogenase, acetyl-CoA acetyltransferase, citrate lyase, and GMP synthase, which are all foundational in essential metabolic processes. (2)
Cell structure and metabolism
4.1 Unique Physiochemical properties The proteome of H. marismortui is highly acidic in order to maintain the structure and function of proteins in a high saline cytoplasm. The acidic residues in these proteins are mostly present on the surface of the folded proteins, making its average isoelectric point a low 5.0. It is expected that having a less negative protein surface charge would cause them to be insoluble in such a high salinity environment. (3)
The major pathways for sugar breakdown in H. marismortui are glycolysis and the modified Entner-Doudoroff (ED) pathways. Energy and amino acid biosynthetic precursors are produced when the products of these pathways are acted upon by the TCA cycle enzymes. The H. marismortui genome sequence codes for enzymes that synthesize up to 16 amino acids. This is more than the standard eight amino acids that are typically coded for in other Halobacterium organisms. Energy and metabolic carbon and nitrogen are also produced via catabolism of amino acids. Inorganic carbons are fixed into sugars via the reductive carboxylate cycle, which uses phosphophenol pyruvate carboxylase and gluconeogensis enzymes. (3, 5)
4.3 Environmental Response System
Due to the high saline, low oxygen solubility, and high light intensity environment, H. marismortui has a complex photobiological response system that includes opsins, cryptochrome/photolyase, clock regulators and transducers. This characteristic is also found in Halobacterium sp. NRC-1. Opsin proteins take advantage of the high light intensity by using light energy to maintain physiological ion concentrations, facilitate phototaxis, and make energy via a proton gradient. There are six opsin genes in H. marismortui.
Transducers are important in communicating an environmental factor to the chemotaxis apparatus and metabolic pathways. That way, an organism is able to be sensitive to its environment and can change in response in order to better survive. H. marismortui encodes 21 proteins that are associated with transmembrane receptor proteins that act as transducers in the cell. (3)
This organism is found in the Dead Sea, a high saline, low oxygen solubility, and high light intensity environment. Other halophilic archaeal organisms live in solar salterns like the Great Salt Lake, which are extreme environments that have a molarity of about 4.5 molar salt. There is not extensive research done on the interactions of this organism with other organisms that live in its environment. There is not extensive research on its contribution to its immediate environment or on its effect on the environment. However, it is known that the environment effects H. marismortui by increasing the salt concentration within its cells. In fact, it is the high saline environment that makes this organism unique. (1)
There are no known diseases that are caused by this organism. It has not been found to be pathogenic.
Application to Biotechnology
7.1 Use in Cataloguing and Recognizing RNA conformational states
The large ribosome subunit of H. marismortui is used to identify and categorize conformational states of RNA, which are then used to investigate statistical laws that direct those states. Torsion matching and binning are two approaches used in order to describe each discreet conformation sate of the large ribosome subunit. This particular subunit is used because of its high resolution due to its crystal structure as well as its large size. Therefore, it is ideal for use as a target in automated pattern-recognition approaches, such as torsion matching and binning. (6)
7.2 Haloalkaliphilic archaea plasmid used to make shuttle between haloarchaea and E. Coli
A plasmid named pNB101 was isolated from a Haloalkaliphilic archaea had ColE1 replicon of E. coli and two antibiotic resistant genes inserted into it. The result was a new shuttle vector between E. coli and haloarchaea named pNB102. PCR, Southern blotting and restriction endonuclease digestion confirmed the presence of the vector in the transformed haloarchaea species. This was one of the first stages in creating a vector/host system in Haloarchaea.
Although the plasmid used in this application is not from H. marismortui, it is significant to this species because the application of it may be used for other Haloarchaea like H. marismortui. In addition, the cells to be transformed in order to see if transformation was successful needed to be in non-alkaliphilic halocarchaea. Therefore, it is possible that H. marismortui could be used in order to complete the experiment. (7)
7.3 Haloarchaea as forerunner in transformation studies
Furthermore, Haloarchaea were the first microbes that could successfully be transformed, meaning they were used extensively for vector studies, reporter genes, and the development of other genetic tools. (8)
8.1 Tolerance of Ultraviolet Light Exposure
The habitat of H. marismortui and other Halobacteriaceae has a high light intensity, making them susceptible to a large amount of ultraviolet (UV) light exposure. UV light exposure, even in limited amounts, chemically changes DNA by reacting with cytosine and a double bond thymine, both found in DNA, and producing pyrimidine dimers and photoproducts. Therefore, research was conducted to investigate how these organisms are able to withstand such extreme UV light exposure and still proliferate normally. It revealed that proteomic structures and genomic features contribute to the organism’s reduction of UV irradiation induced damage. For example, it was found that these organisms may reduce UV damage in its genome by having a low dipyrimidine content in the genome. The organism also minimizes its use of nucleotide residues that are susceptible to the attack of reactive oxygen species, such as free radicals resulting from UV light. Both these features decrease the chances of protein damage. Results from analyzing the putative mismatch pathway via a correlation with zim gene expression and genomic structure showed that this organism is maximized proteomic and genomic structures to protect itself from the possible damage a high UV light intensity environment may cause an organism. (9)
8.2 The Stability of Halophilic Proteins
Studies were done involving malate dehydrogenase, an essential enzyme in the TCA cycle, to investigate how it maintains its stability in the presence of large concentrations of salt. Based on observation, it is known that proteins in environments with large salt concentrations are halophilic, meaning they have developed the ability to be soluble, stable and active in such extreme environments as the Dead Sea, the habitat of the H. marismortui. Using crystallographic studies, it was shown that the surface of halophilic proteins is enriched by acidic amino acids in order to maintain stability in high salt concentrations. Site-directed mutagenesis of tetrameric malate dehydrogenase was also done in order to analyze the significance of chloride-binding sites in the protein. Results showed that mutating the enzyme involved in facilitating the chloride binding site affected the protein’s stability significantly when exposed to KCl, but not KF. Because of the different affects the anions had on the oligomeric states of the mutated enzyme, researchers concluded that the anion-binding sites in halophilic proteins are indeed adapted to the presence of salt in their environment. (10)
8.3 Cell Water Movement within Cell
Neutron scattering using spectrometers were used to investigate the water dynamics inside H. marismortui. Results showed that cell water consisting of 3.5M NaCl solution inside the cells is much slower than water inside cells that do not contain NaCl. About 76% of all cell water in H. marismortui is about 250 times slower than the cells in the control water. Escherichia coli cells were also analyzed using a neutron spectrometer. They did not show the same results as those of H. marismortui. The researchers hypothesize that this unique water movement is due to the presence of a water structure that facilitates the binding of K+ with the cells that have a slower water movement. Further research is needed in order to gain a better understanding about this feature of H. marismortui. (11)
(1) NCBI: Haloarcula marismortui, Accessed June 4, 2007, <http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=2238&lvl=3&p=mapview&p=has_linkout&p=blast_url&p=genome_blast&lin=f&keep=1&srchmode=1&unlock>
(2) Baliga N, Bonneau R, Facciotti M, Pan M, Glusman G, Deutsch E, Shannon P, Chiu Y, Weng R, Gan R, Hung P, Date S, Marcotte E, Hood L, and Ng W. “Genome sequence of Haloarcula marismortui: A halophilic archaeon from the Dead Sea,” Genome Res., Nov 2004; 14: 2221 - 2234.
(3) Kennedy SP, Ng WV, Salzberg SL, Hood L, and DasSarma S. “Understanding the adaptation of Halobacterium species NRC-1 to its extreme environment through computational analysis of its genome sequence,” Genome Res. 2001: 11: 1641–1650.
(4) Oren A, Lau PP, Fox, GE. “The taxonomic status of Halobacterium marismortui from the Dead Sea: a comparison with Halobacterium vallismortis,” Syst Appl Microbiol. 1988;10(3):251-8.
(5) HAMAP: Haloarcula marismortui (Halobacterium marismortui) complete proteome, Accessed May 2, 2007, <http://expasy.org/sprot/hamap/HALMA.html>.
(6) Hershkovitz E, Tannenbaum E, Howerton S, Sheth A, Tannenbaum A, Williams L. “Automated identification of RNA conformation motifs: theory and application to the HM LSU 23S rRNA,” Nucleic Acids Res. 2003 November 1; 31(21): 6249–6257.
(7) Zhou M, Xiang H, Sun C, Tan H. “Construction of a novel shuttle vector based on an RCR-plamid from a haloalkaliphilic archaeon and transformation into other haloarchaea,” Biotechnol Lett. 2004 Jul;26(14):1107-13.
(8) Soppa J. “From genomes to function: haloarchaea as model organisms,” Microbiology 2006, 152: 585-590.
(9) Zhou P, Wen J, Oren A, Chen M, Wu M. “Genomic survey of sequence features for ultraviolet tolerance in haloarchaea (family Halobacteriaceae),” Genomics. 2007 May 9: epublication ahead of print, Accessed June 5, 2007 <http://www.sciencedirect.com/science?_ob=ArticleURL&_udi=B6WG1-4NPHMM9-6&_user=10&_coverDate=05%2F11%2F2007&_rdoc=1&_fmt=&_orig=search&_sort=d&view=c&_acct=C000050221&_version=1&_urlVersion=0&_userid=10&md5=f2af62764e41a5ce9ba2a3e68826d8c7>
(10) Madern D, Ebel C. “Influence of an anion-binding site in the stabilization of halophlic malate dehydrogenase from Haloarcula marismortui,” Biochimie. 2007 May 19: epublication ahead of print, Accessed June 5, 2007 http://www.sciencedirect.com/science?_ob=ArticleListURL&_method=list&_ArticleListID=584987112&_sort=d&_acct=C000050221&_version=1&_urlVersion=0&_userid=10&md5=bda27475ef40d285c5f4759ae521ae75&view=f
(11) Tehei M, Franzetti B, Wood K, Gabel F, Fabiani E, Jasnin M, Zamponi M, Oesterhelt D, Zaccai G, Ginzburg M, Ginzburg BZ. “Neutron scattering reveals extremely slow cell water in a Dead Sea organism,” Proc Natl Acad Sci U S A. 2007 Jan 16;104(3):766-71. Epub 2007 Jan 10, Accessed June 6, 2007 < http://www.pnas.org/cgi/content/abstract/104/3/766>
Edited by Lauren Joe, student of Rachel Larsen and Kit Pogliano