In
materials science and
molecular biology
Molecular biology is the branch of biology that seeks to understand the molecular basis of biological activity in and between cells, including biomolecular synthesis, modification, mechanisms, and interactions. The study of chemical and phys ...
, thermostability is the ability of a
substance to resist irreversible change in its
chemical or physical structure, often by resisting
decomposition or
polymerization, at a high relative
temperature
Temperature is a physical quantity that expresses quantitatively the perceptions of hotness and coldness. Temperature is measured with a thermometer.
Thermometers are calibrated in various temperature scales that historically have relied on ...
.
Thermostable materials may be used industrially as
fire retardant
A fire retardant is a substance that is used to slow down or stop the spread of fire or reduce its intensity. This is commonly accomplished by chemical reactions that reduce the flammability of fuels or delay their combustion. Fire retardants ...
s. A ''thermostable
plastic'', an uncommon and unconventional term, is likely to refer to a
thermosetting plastic that cannot be reshaped when heated, than to a
thermoplastic that can be remelted and recast.
Thermostability is also a property of some
protein
Proteins are large biomolecules and macromolecules that comprise one or more long chains of amino acid residues. Proteins perform a vast array of functions within organisms, including catalysing metabolic reactions, DNA replication, respon ...
s. To be a thermostable protein means to be resistant to changes in
protein structure due to applied heat.
Thermostable proteins
Most life-forms on Earth live at temperatures of less than 50 °C, commonly from 15 to 50 °C. Within these organisms are macromolecules (proteins and nucleic acids) which form the three-dimensional structures essential to their enzymatic activity. Above the native temperature of the organism, thermal energy may cause the
unfolding and
denaturation, as the heat can disrupt the intramolecular bonds in the tertiary and quaternary structure. This unfolding will result in loss in enzymatic activity, which is understandably deleterious to continuing life-functions. An example of such is the denaturing of proteins in
albumen from a clear, nearly colourless liquid to an opaque white, insoluble gel.
Proteins capable of withstanding such high temperatures compared to proteins that cannot, are generally from microorganisms that are hyperthermophiles. Such organisms can withstand above 50 °C temperatures as they usually live within environments of 85 °C and above.
Certain
thermophilic life-forms exist which can withstand temperatures above this, and have corresponding adaptations to preserve protein function at these temperatures. These can include altered bulk properties of the cell to stabilize all proteins, and specific changes to individual proteins. Comparing
homologous
Homology may refer to:
Sciences
Biology
*Homology (biology), any characteristic of biological organisms that is derived from a common ancestor
*Sequence homology, biological homology between DNA, RNA, or protein sequences
* Homologous chrom ...
proteins present in these thermophiles and other organisms reveal some differences in the protein structure. One notable difference is the presence of extra
hydrogen bonds in the thermophile's proteins—meaning that the protein structure is more resistant to unfolding. Similarly, thermostable proteins are rich in
salt bridges or/and extra
disulfide bridges stabilizing the structure. Other factors of protein thermostability are compactness of protein structure, oligomerization, and strength interaction between subunits.
Uses and applications
Polymerase chain reactions
Thermostable
enzyme
Enzymes () are proteins that act as biological catalysts by accelerating chemical reactions. The molecules upon which enzymes may act are called substrate (chemistry), substrates, and the enzyme converts the substrates into different molecule ...
s such as
Taq polymerase and
Pfu DNA polymerase are used in
polymerase chain reactions (PCR) where temperatures of 94 °C or over are used to melt
DNA strands in the denaturation step of PCR. This resistance to high temperature allows for DNA polymerase to elongate DNA with a desired sequence of interest with the presence of dNTPs.
Feed additives
Enzymes are often added to animal feed to improve the health and growth of farmed animals, particularly chickens and pigs. The feed is typically treated with high pressure steam to kill bacteria such as
Salmonella. Therefore the added enzymes (e.g.
phytase and
xylanase) must be able to withstand this thermal challenge without being irreversibly inactivated.
Protein purification
Knowledge of an enzyme's resistance to high temperatures is especially beneficial in
protein purification. In the procedure of heat denaturation, one can subject a mixture of proteins to high temperatures, which will result in the denaturation of proteins that are not thermostable, and the isolation of the protein that is thermodynamically stable. One notable example of this is found in the purification of
alkaline phosphatase from the hyperthermophile ''
Pyrococcus abyssi''. This enzyme is known for being heat stable at temperatures greater than 95 °C, and therefore can be partially purified by heating when heterologously expressed in ''E. coli''. The increase in temperature causes the ''E. coli'' proteins to precipitate, while the ''P. abyssi'' alkaline phosphatase remains stably in solution.
Glycoside hydrolases
Another important group of thermostable enzymes are
glycoside hydrolases. These enzymes are responsible of the degradation of the major fraction of biomass, the polysaccharides present in starch and lignocellulose. Thus,
glycoside hydrolases are gaining great interest in biorefining applications in the future bioeconomy. Some examples are the production of monosaccharides for food applications as well as use as carbon source for microbial conversion in fuels (ethanol) and chemical intermediates, production of oligosaccharides for prebiotic applications and production of surfactants alkyl glycoside type. All of these processes often involve thermal treatments to facilitate the polysaccharide hydrolysis, hence give thermostable variants of
glycoside hydrolases an important role in this context.
Approaches to improve thermostability of proteins
Protein engineering can be used to enhance the thermostability of proteins. A number of
site-directed and
random mutagenesis
In molecular biology, mutagenesis is an important laboratory technique whereby DNA mutations are deliberately engineered to produce libraries of mutant genes, proteins, strains of bacteria, or other genetically modified organisms. The various ...
techniques, in addition to
directed evolution, have been used to increase the thermostability of target proteins. Comparative methods have been used to increase the stability of
mesophilic
A mesophile is an organism that grows best in moderate temperature, neither too hot nor too cold, with an optimum growth range from . The optimum growth temperature for these organisms is 37°C. The term is mainly applied to microorganisms. Org ...
proteins based on comparison to
thermophilic homologs. Additionally, analysis of the protein unfolding by
molecular dynamics
Molecular dynamics (MD) is a computer simulation method for analyzing the physical movements of atoms and molecules. The atoms and molecules are allowed to interact for a fixed period of time, giving a view of the dynamic "evolution" of th ...
can be used to understand the process of unfolding and then design stabilizing mutations. Rational protein engineering for increasing protein thermostability includes mutations which truncate loops, increase salt bridges or hydrogen bonds, introduced
disulfide bonds. In addition, ligand binding can increase the stability of the protein, particularly when purified. There are various different forces that allow for the thermostability of a particular protein. These forces include hydrophobic interactions, electrostatic interactions, and the presence of disulfide bonds. The overall amount of hydrophobicity present in a particular protein is responsible for its thermostability. Another type of force that is responsible for thermostability of a protein is the electrostatic interactions between molecules. These interactions include salt bridges and hydrogen bonds. Salt bridges are unaffected by high temperatures, therefore, are necessary for protein and enzyme stability. A third force used to increase thermostability in proteins and enzymes is the presence of disulfide bonds. They present covalent cross-linkages between the polypeptide chains. These bonds are the strongest because they’re covalent bonds, making them stronger than intermolecular forces.
Glycosylation is another way to improve the thermostability of proteins.
Stereoelectronic effects in stabilizing interactions between carbohydrate and protein can lead to the thermostabilization of the glycosylated protein.
Cyclizing enzymes by covalently linking the N-terminus to the C-terminus has been applied to increase the thermostability of many enzymes.
Intein cyclization and
SpyTag/SpyCatcher cyclization have often been employed.
Thermostable toxins
Certain
poisonous
fungi
A fungus (plural, : fungi or funguses) is any member of the group of Eukaryote, eukaryotic organisms that includes microorganisms such as yeasts and Mold (fungus), molds, as well as the more familiar mushrooms. These organisms are classified ...
contain thermostable
toxins, such as
amatoxin found in the
death cap and
autumn skullcap mushroom
A mushroom or toadstool is the fleshy, spore-bearing fruiting body of a fungus, typically produced above ground, on soil, or on its food source. ''Toadstool'' generally denotes one poisonous to humans.
The standard for the name "mushroom" is ...
s and
patulin from molds. Therefore, applying heat to these will not remove the toxicity and is of particular concern for food safety.
See also
;Thermophiles
*''
Thermus thermophilus
''Thermus thermophilus'' is a Gram-negative bacterium used in a range of biotechnological applications, including as a model organism for genetic manipulation, structural genomics, and systems biology. The bacterium is extremely thermophilic ...
''
*''
Thermus aquaticus''
*''
Pyrococcus furiosus''
References
External links
{{wiktionary
Thermostability of Proteins
Protein structure
Toxicology
Extremophiles