In
materials science and
molecular biology, 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.
Thermostable materials may be used industrially as
fire retardants. A ''thermostable
plastic'', an uncommon and unconventional term, is likely to refer to a
thermosetting plastic
In materials science, a thermosetting polymer, often called a thermoset, is a polymer that is obtained by irreversibly hardening (" curing") a soft solid or viscous liquid prepolymer (resin). Curing is induced by heat or suitable radiation and ...
that cannot be reshaped when heated, than to a
thermoplastic that can be remelted and recast.
Thermostability is also a property of some
proteins. To be a thermostable protein means to be resistant to changes in
protein structure
Protein structure is the three-dimensional arrangement of atoms in an amino acid-chain molecule. Proteins are polymers specifically polypeptides formed from sequences of amino acids, the monomers of the polymer. A single amino acid monomer ma ...
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
Egg white is the clear liquid (also called the albumen or the glair/glaire) contained within an egg. In chickens it is formed from the layers of secretions of the anterior section of the hen's oviduct during the passage of the egg. It forms arou ...
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 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
In biochemistry, a disulfide (or disulphide in British English) refers to a functional group with the structure . The linkage is also called an SS-bond or sometimes a disulfide bridge and is usually derived by the coupling of two thiol groups. In ...
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
enzymes such as
Taq polymerase and
Pfu DNA polymerase are used in
polymerase chain reaction
The polymerase chain reaction (PCR) is a method widely used to rapidly make millions to billions of copies (complete or partial) of a specific DNA sample, allowing scientists to take a very small sample of DNA and amplify it (or a part of it) ...
s (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
Endo-1,4-β-xylanase (EC 3.2.1.8, systematic name 4-β-D-xylan xylanohydrolase) is any of a class of enzymes that degrade the linear polysaccharide xylan into xylose, thus breaking down hemicellulose, one of the major components of plant cel ...
) 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 Protein purification is a series of processes intended to isolate one or a few proteins from a complex mixture, usually cells, tissues or whole organisms. Protein purification is vital for the specification of the function, structure and interact ...
. 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
The enzyme alkaline phosphatase (EC 3.1.3.1, alkaline phosphomonoesterase; phosphomonoesterase; glycerophosphatase; alkaline phosphohydrolase; alkaline phenyl phosphatase; orthophosphoric-monoester phosphohydrolase (alkaline optimum), systematic ...
from the hyperthermophile ''
Pyrococcus abyssi
''Pyrococcus abyssi'' is a hyperthermophilic archaeon isolated from a deep-sea hydrothermal vent in the North Fiji Basin at . It is anaerobic, sulfur-metabolizing, gram-negative, coccus-shaped and highly motile. Its optimum growth temperature ...
''. 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
Glycoside hydrolases (also called glycosidases or glycosyl hydrolases) catalyze the hydrolysis of glycosidic bonds in complex sugars. They are extremely common enzymes with roles in nature including degradation of biomass such as cellulose (cel ...
. These enzymes are responsible of the degradation of the major fraction of biomass, the polysaccharides present in starch and lignocellulose. Thus,
glycoside hydrolases
Glycoside hydrolases (also called glycosidases or glycosyl hydrolases) catalyze the hydrolysis of glycosidic bonds in complex sugars. They are extremely common enzymes with roles in nature including degradation of biomass such as cellulose (cel ...
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
Glycoside hydrolases (also called glycosidases or glycosyl hydrolases) catalyze the hydrolysis of glycosidic bonds in complex sugars. They are extremely common enzymes with roles in nature including degradation of biomass such as cellulose (cel ...
an important role in this context.
Approaches to improve thermostability of proteins
Protein engineering
Protein engineering is the process of developing useful or valuable proteins. It is a young discipline, with much research taking place into the understanding of protein folding and recognition for protein design principles. It has been used to im ...
can be used to enhance the thermostability of proteins. A number of
site-directed and
random mutagenesis techniques, in addition to
directed evolution
Directed evolution (DE) is a method used in protein engineering that mimics the process of natural selection to steer proteins or nucleic acids toward a user-defined goal. It consists of subjecting a gene to iterative rounds of mutagenesis ...
, 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
A thermophile is an organism—a type of extremophile—that thrives at relatively high temperatures, between . Many thermophiles are archaea, though they can be bacteria or fungi. Thermophilic eubacteria are suggested to have been among the ea ...
homologs. Additionally, analysis of the protein unfolding by
molecular dynamics 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 bond
In biochemistry, a disulfide (or disulphide in British English) refers to a functional group with the structure . The linkage is also called an SS-bond or sometimes a disulfide bridge and is usually derived by the coupling of two thiol groups. In ...
s. 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
Glycosylation is the reaction in which a carbohydrate (or 'glycan'), i.e. a glycosyl donor, is attached to a hydroxyl or other functional group of another molecule (a glycosyl acceptor) in order to form a glycoconjugate. In biology (but not a ...
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 contain thermostable
toxins, such as
amatoxin Amatoxin is the collective name of a subgroup of at least nine related toxic compounds found in three genera of poisonous mushrooms ('' Amanita'', ''Galerina'' and '' Lepiota'') and one species ( Conocybe filaris) of the genus '' Conocybe''. Amatoxi ...
found in the
death cap
''Amanita phalloides'' (), commonly known as the death cap, is a deadly poisonous basidiomycete fungus, one of many in the genus ''Amanita''. Widely distributed across Europe, but now sprouting in other parts of the world, ''A. phalloid ...
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 t ...
s and
patulin
Patulin is an organic compound classified as a polyketide. It is a white powder soluble in acidic water and in organic solvents. It is a lactone that is heat-stable, so it is not destroyed by pasteurization or thermal denaturation.http://www.sig ...
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 aquaticus''
*''
Pyrococcus furiosus''
References
External links
{{wiktionary
Thermostability of Proteins
Protein structure
Toxicology
Extremophiles