Extremophile & Adaptation
-
There are several of the existing MDH clones expressed in organisms that have unique “extremophile” characteristics. Extremophiles are organisms that thrive in extreme environments unsuitable for most life forms. They exist in diverse habitats on Earth, such as deep-sea hydrothermal vents, polar ice caps, hot springs, and volcanic regions. There are various types of extremophiles. Thermophiles thrive in very high temperatures, while psychrophiles adapt to extremely cold conditions. Acidophiles survive in highly acidic environments, alkaliphiles tolerate highly alkaline conditions, and halophiles can live in environments with high salt concentrations. Barophiles or piezophiles withstand high pressures. Proteins in extremophiles have evolved specific adaptations to function and maintain stability in extreme conditions. By employing strategies such as increased stability, flexibility, and surface properties, these proteins enable organisms to thrive in extreme temperature, cold, or high-salt conditions.
Thermophilic Proteins: Proteins from thermophilic organisms that thrive in high-temperature environments have enhanced stability. They often possess increased numbers of salt bridges, stronger hydrogen bonding networks, and more hydrophobic interactions. These features help maintain protein structure and prevent unfolding or denaturation at elevated temperatures. Additionally, thermophilic proteins exhibit higher structural rigidity and packing density, enabling them to withstand the heat.
Psychrophilic Proteins: Proteins from psychrophiles adapted to cold environments have unique adaptations to remain functional at low temperatures. They often exhibit increased flexibility and surface mobility, allowing for enzyme-substrate interactions at lower energy levels. Psychrophilic proteins may also have a higher content of alpha-helices, which enhances their flexibility and facilitates enzymatic activity in cold conditions.
Halophilic Proteins: Proteins from halophiles, which inhabit high-salt environments, possess adaptations to cope with osmotic stress. They typically have a higher proportion of negatively charged amino acids on their surfaces. This enables interactions and binding with water molecules, maintaining protein structure and function in dehydrating conditions. Additionally, halophilic proteins may exhibit increased hydrophobicity and optimized interdomain interactions to withstand the high salt concentrations.
-
The diverse array of extremophile proteins, with their unique properties and adaptations, holds great potential for advancements in biotechnology, industrial processes, environmental remediation, and healthcare. These proteins serve as valuable resources for innovation, providing opportunities for novel applications and addressing challenges in a wide range of scientific and technological fields. Proteins from extremophiles, including thermophiles, psychrophiles, and halophiles, have found applications in various fields such as biotechnology, engineering, and medicine. Here are some examples:
Thermostable enzymes from thermophiles, such as DNA polymerases and amylases, are widely used in polymerase chain reaction (PCR) and industrial processes that require high-temperature conditions. Psychrophilic enzymes have applications in low-temperature biotechnological processes, such as cold-active amylases used in the food industry and detergents. Cold-adapted enzymes have also been used for application involving biofuel production, waste treatment, and detergent formulation. In medicine, enzymes with cold-adapted characteristics are being explored for potential medical applications, including cryosurgery, preservation of tissues, and development of cold-active drugs. Halophilic enzymes, such as halolysins and halophilic proteases, have been explored for their potential in biocatalysis and bioremediation. Halophilic proteins have shown promise in various medical fields, including the development of osmoprotectants for cell preservation, wound healing agents, and antimicrobial peptides. Acidophilic enzymes, such as acid proteases and acid lipases, have found utility in the food industry for enhancing specific flavors and textures. These proteins contribute to the production of various food products and help create unique sensory experiences. Alkaliphilic enzymes, including alkaline proteases and alkaline phosphatases, have proven valuable in detergent formulations, where they assist in breaking down stubborn stains and improving cleaning efficiency. Additionally, these enzymes are utilized in bioremediation processes, aiding in the degradation of pollutants under alkaline conditions.
There are several MDH clones from extremophiles whose properties are not fully investigated. When coupled with a non-extremophilic MDH clone, a number of interesting projects could arise.
Information & Resources
Chlorobaculum parvum MDH_CHLP8: Chlorobaculum parvum (isolated from bacterial strain NCIB 8327), formerly Chlorobium vibrioforme, is a green sulfur photosynthetic bacterium which contains bacteriochlorophyl D and the caratenoid chlorobactene. This organism grows photolithotrophically using sulfide, sulfur or thiosulfate as electron donors. Heterotetramer (dimer of dimers). Very little is published on this enzyme.
Streptomyces avermitilis MDH_STRAW: Bacteria first isolated in 1979 at Kitasato Institute from a soil sample collected at Kawana, Ito City, Shizuoka Prefecture, Japan.This particular Streptomyces species dwells in terrestrial soils and has a brownish-gray spore mass. The spores are spherical (as opposed to oval) with a smooth spore surface and come in chains of more than 15. The sporophores form spiral side branches on aerial mycelia. S. avermitilis is an important species to have its genome sequenced because it produces certain secondary metabolites, called avermectins, that have antihelmintic and insecticidal properties. This is a thermostable MDH with a duel NADH and NADPH substrate. Strongly inhibited by both OAA and NADH, while other ions increase activity.
Haloarcula marismortui. DH_HALMA: 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, thrives in this extreme environment due to several adaptations in protein structure, metabolic strategies and physiologic responses. The quaternary structure is stabilized by chloride ions bound between the subunits. This may be an adaptation to the halophilic environment. Sequence is similar to LDH, more than other MDH isoforms. Most haliophiles are more active in high salt concentrations.
Aquaspirillum arcticum MDH_AQUAR: Bactrium grown in sediment under snow and ice, in the northwest territory of Canada. Produces cold-shock proteins and cold-acclimation proteins. Even though most cold tolerant proteins are more flexible than thermostable proteins, the sequence of this MDH is highly related to thermophilic MDH. Yet the half-life at high temps for the two proteins are very different. This protein is 2-3 fold more active at 4-10 oC than other MDH enzymes – possibly due to an increased flexibility at the active site as well as possible reduced interactions at subunits. This may be an interesting enzyme to look for regulation between subunits.
Ignicoccus islandicus A0A0U3FQH7_9CREN aka 9CREN. Originally isolated from Ignicoccus islandicus DSM 13165, a genus of Archaea living in a hydrothermal vent growing in ~90oC. This enzyme has an NADPH dependent, tetrameric with LDH like activity. MDH from this organism may be a divergent homolog from MDH and LDH with different allosteric properties.
Relevant Publications
Anoxygenic Phototrophic Bacteria from Extreme Environments (Imhoff 2017).
Malate dehydrogenase from Chlorobium vibrioforme, Chlorobium tepidum, and Heliobacterium gestii: purification, characterization, and investigation of dinucleotide binding by dehydrogenases by use of empirical methods of protein sequence analysis (J Bacterial 1992)
Expression and identification of a thermostable malate dehydrogenase from multicellular prokaryote Streptomyces avermitilis MA-4680 (Mol Biol Rep, 2011, 38: 1629-36)
Characterization of malate dehydrogenase from the hyperthermophilic archaeon Pyrobaculum islandicum (Extremophiles 2007)
Identification and biochemical characterization of a thermostable malate dehydrogenase from the mesophile Streptomyces coelicolor A3(2). (Biosci Biotechnol Biochem 2010)
Cloning, sequencing, and expression in Escherichia coli of the gene coding for malate dehydrogenase of the extremely halophilic archaebacterium Haloarcula marismortui (Biochemistry, 1993 32:4308-13)
Structural basis for cold adaptation. Sequence, biochemical properties, and crystal structure of malate dehydrogenase from a psychrophile Aquaspirillium arcticum (JBC 1999, 274: 11761-7)
The archaeal LDH-like malate dehydrogenase from Ignicoccus islandicus displays dual substrate recognition, hidden allostery and a non-canonical tetrameric oligomeric organization (J Struct Biol 2019)
List of Skills/ Techniques Needed: Basic molecular techniques for SDM and protein expression/purification. Thermal stability (both activity and folding) and other biophysical approaches.
A list of adopters:: Jessica Bell, Ellis Bell, Amy Springer…. (email josephprovost@sandiego.edu to add your name!)
