Cellulase (EC 3.2.1.4; systematic name 4-β-
D-glucan 4-glucanohydrolase) is any of several
enzymes
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 ...
produced chiefly by
fungi
A fungus ( : fungi or funguses) is any member of the group of eukaryotic organisms that includes microorganisms such as yeasts and molds, as well as the more familiar mushrooms. These organisms are classified as a kingdom, separately from ...
,
bacteria
Bacteria (; singular: bacterium) are ubiquitous, mostly free-living organisms often consisting of one biological cell. They constitute a large domain of prokaryotic microorganisms. Typically a few micrometres in length, bacteria were among ...
, and
protozoan
Protozoa (singular: protozoan or protozoon; alternative plural: protozoans) are a group of single-celled eukaryotes, either free-living or parasitic, that feed on organic matter such as other microorganisms or organic tissues and debris. Histo ...
s that
catalyze
Catalysis () is the process of increasing the rate of a chemical reaction by adding a substance known as a catalyst (). Catalysts are not consumed in the reaction and remain unchanged after it. If the reaction is rapid and the catalyst recyc ...
cellulolysis
Cellulose is an organic compound with the formula , a polysaccharide consisting of a linear chain of several hundred to many thousands of β(1→4) linked D-glucose units. Cellulose is an important structural component of the primary cell wall ...
, the decomposition of
cellulose
Cellulose is an organic compound with the formula , a polysaccharide consisting of a linear chain of several hundred to many thousands of β(1→4) linked D-glucose units. Cellulose is an important structural component of the primary cell wall ...
and of some related
polysaccharide
Polysaccharides (), or polycarbohydrates, are the most abundant carbohydrates found in food. They are long chain polymeric carbohydrates composed of monosaccharide units bound together by glycosidic linkages. This carbohydrate can react with wa ...
s:
: Endohydrolysis of (1→4)-β-
D-glucosidic linkages in cellulose, lichenin and cereal β-
D-glucan
The name is also used for any naturally occurring mixture or complex of various such enzymes, that act serially or synergistically to decompose cellulosic material.
Cellulases break down the cellulose molecule into
monosaccharide
Monosaccharides (from Greek ''monos'': single, '' sacchar'': sugar), also called simple sugars, are the simplest forms of sugar and the most basic units (monomers) from which all carbohydrates are built.
They are usually colorless, water-solub ...
s ("simple sugars") such as β-
glucose
Glucose is a simple sugar with the molecular formula . Glucose is overall the most abundant monosaccharide, a subcategory of carbohydrates. Glucose is mainly made by plants and most algae during photosynthesis from water and carbon dioxide, using ...
, or shorter polysaccharides and
oligosaccharide
An oligosaccharide (/ˌɑlɪgoʊˈsækəˌɹaɪd/; from the Greek ὀλίγος ''olígos'', "a few", and σάκχαρ ''sácchar'', "sugar") is a saccharide polymer containing a small number (typically two to ten) of monosaccharides (simple sugar ...
s. Cellulose breakdown is of considerable economic importance, because it makes a major constituent of plants available for consumption and use in chemical reactions. The specific reaction involved is the
hydrolysis
Hydrolysis (; ) is any chemical reaction in which a molecule of water breaks one or more chemical bonds. The term is used broadly for substitution reaction, substitution, elimination reaction, elimination, and solvation reactions in which water ...
of the 1,4-β-
D-
glycosidic linkage
A glycosidic bond or glycosidic linkage is a type of covalent bond that joins a carbohydrate (sugar) molecule to another group, which may or may not be another carbohydrate.
A glycosidic bond is formed between the hemiacetal or hemiketal grou ...
s in cellulose,
hemicellulose
A hemicellulose (also known as polyose) is one of a number of heteropolymer, heteropolymers (matrix polysaccharides), such as arabinoxylans, present along with cellulose in almost all embryophyte, terrestrial plant cell walls.Scheller HV, Ulvskov H ...
,
lichenin
Lichenin, also known as lichenan or moss starch, is a complex glucan occurring in certain species of lichens. It can be extracted from ''Cetraria islandica'' (Iceland moss). It has been studied since about 1957.
Structure
Chemically, lichenin i ...
, and cereal
β-D-glucans. Because cellulose molecules bind strongly to each other, cellulolysis is relatively difficult compared to the breakdown of other polysaccharides such as starch.
Most mammals have only very limited ability to digest dietary fibres like cellulose by themselves. In many herbivorous animals such as
ruminant
Ruminants (suborder Ruminantia) are ungulate, hoofed herbivorous grazing or browsing mammals that are able to acquire nutrients from plant-based food by Enteric fermentation, fermenting it in a specialized stomach prior to digestion, principally ...
s like cattle and sheep and
hindgut fermenters
Hindgut fermentation is a digestive process seen in monogastric herbivores, animals with a simple, single-chambered stomach. Cellulose is digested with the aid of symbiotic bacteria.[symbiotic
Symbiosis (from Greek , , "living together", from , , "together", and , bíōsis, "living") is any type of a close and long-term biological interaction between two different biological organisms, be it mutualistic, commensalistic, or parasit ...]
bacteria. Endogenous cellulases are produced by a few types of
metazoan
Animals are multicellular, eukaryotic organisms in the biological kingdom Animalia. With few exceptions, animals Heterotroph, consume organic material, Cellular respiration#Aerobic respiration, breathe oxygen, are Motility, able to move, ca ...
animals, such as some
termite
Termites are small insects that live in colonies and have distinct castes (eusocial) and feed on wood or other dead plant matter. Termites comprise the infraorder Isoptera, or alternatively the epifamily Termitoidae, within the order Blattode ...
s, snails,
and
earthworm
An earthworm is a terrestrial invertebrate that belongs to the phylum Annelida. They exhibit a tube-within-a-tube body plan; they are externally segmented with corresponding internal segmentation; and they usually have setae on all segments. Th ...
s.
Recently, cellulases have also been found in green microalgae (''Chlamydomonas reinhardtii'', ''Gonium pectorale'' and ''Volvox carteri'') and their catalytic domains (CD) belonging to
GH9 Family show highest
sequence homology
Sequence homology is the biological homology between DNA, RNA, or protein sequences, defined in terms of shared ancestry in the evolutionary history of life. Two segments of DNA can have shared ancestry because of three phenomena: either a spe ...
to metazoan endogenous cellulases. Algal cellulases are modular, consisting of putative novel cysteine-rich
carbohydrate-binding modules (CBMs), proline/serine-(PS) rich linkers in addition to putative Ig-like and unknown domains in some members. Cellulase from ''Gonium pectorale'' consisted of two CDs separated by linkers and with a C-terminal CBM.
[Guerriero G, Sergeant K, Legay S. Hausman J-F, Cauchie H-M, Ahmad I, Siddiqui KS. 2018 Novel insights from comparative in silico analysis of green microalgae cellulases. Int. J. Mol. Sci. 19 (6), 1782.]
Several different kinds of cellulases are known, which differ structurally and mechanistically. Synonyms, derivatives, and specific enzymes associated with the name "cellulase" include
endo-1,4-β-D-glucanase (β-1,4-glucanase, β-1,4-endoglucan hydrolase, endoglucanase D, 1,4-(1,3;1,4)-β-
D-glucan 4-glucanohydrolase),
carboxymethyl cellulase (CMCase), avicelase,
celludextrinase
Cellulase (EC 3.2.1.4; systematic name 4-β-D-glucan 4-glucanohydrolase) is any of several enzymes produced chiefly by fungi, bacteria, and protozoans that catalyze cellulolysis, the decomposition of cellulose and of some related polysaccharid ...
,
cellulase A
Cellulase (EC 3.2.1.4; systematic name 4-β-D-glucan 4-glucanohydrolase) is any of several enzymes produced chiefly by fungi, bacteria, and protozoans that catalyze cellulolysis, the decomposition of cellulose and of some related polysaccharid ...
,
cellulosin AP
Cellulase (EC 3.2.1.4; systematic name 4-β-D-glucan 4-glucanohydrolase) is any of several enzymes produced chiefly by fungi, bacteria, and protozoans that catalyze cellulolysis, the decomposition of cellulose and of some related polysaccharid ...
,
alkali cellulase
Cellulase (EC 3.2.1.4; systematic name 4-β-D-glucan 4-glucanohydrolase) is any of several enzymes produced chiefly by fungi, bacteria, and protozoans that catalyze cellulolysis, the decomposition of cellulose and of some related polysaccharid ...
,
cellulase A 3
Cellulase (EC 3.2.1.4; systematic name 4-β-D-glucan 4-glucanohydrolase) is any of several enzymes produced chiefly by fungi, bacteria, and protozoans that catalyze cellulolysis, the decomposition of cellulose and of some related polysaccharide ...
,
9.5 cellulase, and
pancellase SS. Enzymes that cleave
lignin
Lignin is a class of complex organic polymers that form key structural materials in the support tissues of most plants. Lignins are particularly important in the formation of cell walls, especially in wood and bark, because they lend rigidity ...
have occasionally been called cellulases, but this old usage is deprecated; they are
lignin-modifying enzyme Lignin-modifying enzymes (LMEs) are various types of enzymes produced by fungi and bacteria that catalyze the breakdown of lignin, a biopolymer commonly found in the cell walls of plants. The terms ligninases and lignases are older names for the sam ...
s.
Types and action
Five general types of cellulases based on the type of reaction catalyzed:
*
Endocellulases (EC 3.2.1.4) randomly cleave internal bonds at amorphous sites that create new chain ends.
*
Exocellulase
Glucan 1,4-beta-glucosidase (or 4-beta-D-glucan glucohydrolase) is an enzyme that catalysis, catalyses the hydrolysis of (1->4)-linkages in glucan, 1,4-beta-D-glucans and related oligosaccharides, removing successive glucose units.
This is one of ...
s or cellobiohydrolases (EC 3.2.1.91) cleave two to four units from the ends of the exposed chains produced by endocellulase, resulting in
tetrasaccharide
A tetrasaccharide is a carbohydrate which gives upon hydrolysis four molecules of the same or different monosaccharides. For example, stachyose upon hydrolysis gives one molecule each of glucose and fructose and two molecules of galactose
Gala ...
s
or
disaccharide
A disaccharide (also called a double sugar or ''biose'') is the sugar formed when two monosaccharides are joined by glycosidic linkage. Like monosaccharides, disaccharides are simple sugars soluble in water. Three common examples are sucrose, lact ...
s, such as
cellobiose
Cellobiose is a disaccharide with the formula (C6H7(OH)4O)2O. It is classified as a reducing sugar. In terms of its chemical structure, it is derived from the condensation of a pair of β-glucose molecules forming a β(1→4) bond. It can be hyd ...
. Exocellulases are further classified into type I, that work processively from the reducing end of the cellulose chain, and type II, that work processively from the nonreducing end.
* Cellobiases (EC 3.2.1.21) or
β-glucosidase
β-Glucosidase (EC 3.2.1.21; systematic name β-D-glucoside glucohydrolase) is an enzyme that catalyses the following reaction:
: Hydrolysis of terminal, non-reducing β-D-glucosyl residues with release of β-D-glucose
Structure
β-Glucosidase ...
s hydrolyse the exocellulase product into individual monosaccharides.
*
Oxidative cellulase
Redox (reduction–oxidation, , ) is a type of chemical reaction in which the oxidation states of substrate change. Oxidation is the loss of electrons or an increase in the oxidation state, while reduction is the gain of electrons or a d ...
s depolymerize cellulose by radical reactions, as for instance
cellobiose dehydrogenase (acceptor).
*
Cellulose phosphorylase
Cellulose is an organic compound with the chemical formula, formula , a polysaccharide consisting of a linear chain of several hundred to many thousands of glycosidic bond, β(1→4) linked glucose, D-glucose units. Cellulose is an important stru ...
s depolymerize cellulose using phosphates instead of water.
Avicelase has almost exclusively exo-cellulase activity, since avicel is a highly micro-crystalline substrate.
Within the above types there are also progressive (also known as processive) and nonprogressive types. Progressive cellulase will continue to interact with a single polysaccharide strand, nonprogressive cellulase will interact once then disengage and engage another polysaccharide strand.
Cellulase action is considered to be synergistic as all three classes of cellulase can yield much more sugar than the addition of all three separately. Aside from ruminants, most animals (including humans) do not produce cellulase in their bodies and can only partially break down cellulose through fermentation, limiting their ability to use
energy
In physics, energy (from Ancient Greek: ἐνέργεια, ''enérgeia'', “activity”) is the quantitative property that is transferred to a body or to a physical system, recognizable in the performance of work and in the form of heat a ...
in fibrous plant material.
Structure
Most fungal cellulases have a two-domain structure, with one catalytic domain and one cellulose binding domain, that are connected by a flexible linker. This structure is adapted for working on an insoluble substrate, and it allows the enzyme to diffuse two-dimensionally on a surface in a caterpillar-like fashion. However, there are also cellulases (mostly endoglucanases) that lack cellulose binding domains.
Both binding of substrates and catalysis depend on the three-dimensional structure of the enzyme which arises as a consequence of the level of
protein folding
Protein folding is the physical process by which a protein chain is translated to its native three-dimensional structure, typically a "folded" conformation by which the protein becomes biologically functional. Via an expeditious and reproduci ...
. The amino acid sequence and arrangement of their residues that occur within the active site, the position where the substrate binds, may influence factors like binding affinity of ligands, stabilization of substrates within the active site and catalysis. The substrate structure is complementary to the precise active site structure of enzyme. Changes in the position of residues may result in distortion of one or more of these interactions. Additional factors like temperature, pH and metal ions influence the non-covalent interactions between enzyme structure. The ''Thermotoga maritima'' species make cellulases consisting of 2 β-sheets (protein structures) surrounding a central catalytic region which is the active-site.
The enzyme is categorised as an endoglucanase, which internally cleaves β-1,4-glycosydic bonds in cellulose chains facilitating further degradation of the polymer. Different species in the same family as ''T. maritima'' make cellulases with different structures.
Cellulases produced by the species ''Coprinopsis cinerea'' consists of seven protein strands in the shape of an enclosed tunnel called a β/α barrel. These enzymes hydrolyse the substrate carboxymethyl cellulose. Binding of the substrate in the active site induces a change in conformation which allows degradation of the molecule.
Cellulase complexes
In many bacteria, cellulases in vivo are complex enzyme structures organized in
supramolecular complexes, the
cellulosomes
Cellulosomes are multi-enzyme extracellular complexes. Cellulosomes are associated with the cell surface and mediate cell attachment to insoluble substrates and degrade them to soluble products which are then absorbed. Cellulosome complexes are in ...
. They can contain, but are not limited to, five different enzymatic subunits representing namely endocellulases, exocellulases, cellobiases, oxidative cellulases and cellulose phosphorylases wherein only exocellulases and cellobiases participate in the actual hydrolysis of the β(1→4) linkage. The number of sub-units making up cellulosomes can also determine the rate of enzyme activity.
Multidomain cellulases are widespread among many taxonomic groups, however, cellulases from anaerobic bacteria, found in cellulosomes, have the most complex architecture consisting of different types of modules. For example, ''Clostridium cellulolyticum'' produces 13 GH9 modular cellulases containing a different number and arrangement of catalytic-domain (CD), carbohydrate-binding module (CBM), dockerin, linker and Ig-like domain.
The cellulase complex from ''
Trichoderma reesei
''Trichoderma reesei'' is a mesophilic and filamentous fungus. It is an anamorph of the fungus ''Hypocrea jecorina''.
''T. reesei'' can secrete large amounts of cellulolytic enzymes (cellulases and hemicellulases). Microbial cellulases have ...
'', for example, comprises a component labeled C1 (57,000
dalton
Dalton may refer to:
Science
* Dalton (crater), a lunar crater
* Dalton (program), chemistry software
* Dalton (unit) (Da), the atomic mass unit
* John Dalton, chemist, physicist and meteorologist
Entertainment
* Dalton (Buffyverse), minor cha ...
s) that separates the chains of crystalline cellulose, an endoglucanase (about 52,000 daltons), an exoglucanase (about 61,000 dalton), and a β-glucosidase (76,000 daltons).
[Worthington Biochemical Corporation (2014)]
Cellulase
Accessed on 2014-07-03
Numerous "signature" sequences known as
dockerin
Dockerin is a protein domain found in the cellulosome cellular structure of anaerobic bacteria. It is found on many endoglucanase enzymes. The dockerin's binding partner is the cohesin domain, located on the scaffoldin protein. This interaction ...
s and
cohesin
Cohesin is a protein complex that mediates sister chromatid cohesion, homologous recombination, and DNA looping. Cohesin is formed of SMC3, SMC1, SCC1 and SCC3 ( SA1 or SA2 in humans). Cohesin holds sister chromatids together after DNA rep ...
s have been identified in the
genome
In the fields of molecular biology and genetics, a genome is all the genetic information of an organism. It consists of nucleotide sequences of DNA (or RNA in RNA viruses). The nuclear genome includes protein-coding genes and non-coding ge ...
s of bacteria that produce cellulosomes. Depending on their
amino acid sequence
Protein primary structure is the linear sequence of amino acids in a peptide or protein. By convention, the primary structure of a protein is reported starting from the amino-terminal (N) end to the carboxyl-terminal (C) end. Protein biosynthes ...
and
tertiary structures
Biomolecular structure is the intricate folded, three-dimensional shape that is formed by a molecule of protein, DNA, or RNA, and that is important to its function. The structure of these molecules may be considered at any of several length sc ...
, cellulases are divided into clans and families.
Multimodular cellulases are more efficient than free enzyme (with only CD) due to synergism because of the close proximity between the enzyme and the cellulosic substrate. CBM are involved in binding of cellulose whereas glycosylated linkers provide flexibility to the CD for higher activity and protease protection, as well as increased binding to the cellulose surface.
Mechanism of cellulolysis
Uses
Cellulase is used for commercial food processing in
coffee
Coffee is a drink prepared from roasted coffee beans. Darkly colored, bitter, and slightly acidic, coffee has a stimulant, stimulating effect on humans, primarily due to its caffeine content. It is the most popular hot drink in the world.
S ...
. It performs
hydrolysis
Hydrolysis (; ) is any chemical reaction in which a molecule of water breaks one or more chemical bonds. The term is used broadly for substitution reaction, substitution, elimination reaction, elimination, and solvation reactions in which water ...
of cellulose during drying of
beans
A bean is the seed of several plants in the family Fabaceae, which are used as vegetables for human or animal food. They can be cooked in many different ways, including boiling, frying, and baking, and are used in many traditional dishes th ...
. Furthermore, cellulases are widely used in textile industry and in laundry detergents. They have also been used in the
pulp and paper industry
The pulp and paper industry comprises companies that use wood as raw material and produce pulp, paper, paperboard and other cellulose-based products.
Manufacturing process
The pulp is fed to a paper machine where it is formed as a paper web an ...
for various purposes, and they are even used for pharmaceutical applications.
Cellulase is used in the fermentation of
biomass
Biomass is plant-based material used as a fuel for heat or electricity production. It can be in the form of wood, wood residues, energy crops, agricultural residues, and waste from industry, farms, and households. Some people use the terms bi ...
into
biofuels
Biofuel is a fuel that is produced over a short time span from biomass, rather than by the very slow natural processes involved in the formation of fossil fuels, such as oil. According to the United States Energy Information Administration (E ...
, although this process is relatively experimental at present. Cellulase is used in medicine as a treatment for phytobezoars, a form of cellulose
bezoar
A bezoar is a mass often found trapped in the gastrointestinal system, though it can occur in other locations. A pseudobezoar is an indigestible object introduced intentionally into the digestive system.
There are several varieties of bezoar, s ...
found in the human
stomach
The stomach is a muscular, hollow organ in the gastrointestinal tract of humans and many other animals, including several invertebrates. The stomach has a dilated structure and functions as a vital organ in the digestive system. The stomach i ...
, and it has exhibited efficacy in degrading polymicrobial bacterial
biofilm
A biofilm comprises any syntrophic consortium of microorganisms in which cells stick to each other and often also to a surface. These adherent cells become embedded within a slimy extracellular matrix that is composed of extracellular ...
s by hydrolyzing the β(1-4) glycosidic linkages within the structural, matrix exopolysaccharides of the
extracellular polymeric substance
Extracellular polymeric substances (EPSs) are natural polymers of high molecular weight secreted by microorganisms into their environment. EPSs establish the functional and structural integrity of biofilms, and are considered the fundamental comp ...
(EPS).
Measurement
As the native substrate,
cellulose
Cellulose is an organic compound with the formula , a polysaccharide consisting of a linear chain of several hundred to many thousands of β(1→4) linked D-glucose units. Cellulose is an important structural component of the primary cell wall ...
, is a water-insoluble polymer, traditional reducing sugar assays using this substrate can not be employed for the measurement of cellulase activity. Analytical scientists have developed a number of alternative methods.
* DNSA Method Cellulase activity was determined by incubating 0.5 ml of supernatant with 0.5 ml of 1% carboxymethylcellulose (CMC) in 0.05M citrate buffer (pH 4.8) at 50°C for 30 minutes. The reaction was terminated by the addition of 3 ml dinitrosalicylic acid reagent. Absorbance was read at 540 nm.
A
viscometer
A viscometer (also called viscosimeter) is an instrument used to measure the viscosity of a fluid. For liquids with viscosities which vary with flow conditions, an instrument called a rheometer is used. Thus, a rheometer can be considered as a spec ...
can be used to measure the decrease in viscosity of a solution containing a water-soluble cellulose derivative such as
carboxymethyl cellulose
Carboxymethyl cellulose (CMC) or cellulose gum is a cellulose derivative with carboxymethyl groups (-CH2-COOH) bound to some of the hydroxyl groups of the glucopyranose monomers that make up the cellulose backbone. It is often used as its sodiu ...
upon incubation with a cellulase sample. The decrease in viscosity is directly proportional to the cellulase activity. While such assays are very sensitive and specific for ''endo''-cellulase (''exo''-acting cellulase enzymes produce little or no change in viscosity), they are limited by the fact that it is hard to define activity in conventional enzyme units (micromoles of substrate hydrolyzed or product produced per minute).
Cellooligosaccharide substrates
The lower DP cello-oligosaccharides (DP2-6) are sufficiently soluble in water to act as viable substrates for cellulase enzymes. However, as these substrates are themselves '
reducing sugars
A reducing sugar is any sugar that is capable of acting as a reducing agent. In an alkaline solution, a reducing sugar forms some aldehyde or ketone, which allows it to act as a reducing agent, for example in Benedict's reagent. In such a reactio ...
', they are not suitable for use in traditional reducing sugar assays because they generate a high 'blank' value. However their cellulase mediated hydrolysis can be monitored by
HPLC
High-performance liquid chromatography (HPLC), formerly referred to as high-pressure liquid chromatography, is a technique in analytical chemistry used to separate, identify, and quantify each component in a mixture. It relies on pumps to pa ...
or
IC methods to gain valuable information on the substrate requirements of a particular cellulase enzyme.
Reduced cello-oligosaccharide substrates
Cello-oligosaccharides can be chemically reduced through the action of
sodium borohydride
Sodium borohydride, also known as sodium tetrahydridoborate and sodium tetrahydroborate, is an inorganic compound with the formula Na BH4. This white solid, usually encountered as an aqueous basic solution, is a reducing agent that finds appli ...
to produce their corresponding
sugar alcohols
Sugar alcohols (also called polyhydric alcohols, polyalcohols, alditols or glycitols) are organic compounds, typically derivative (chemistry), derived from sugars, containing one hydroxyl group (–OH) attached to each carbon atom. They are whit ...
. These compounds do not react in reducing sugar assays but their hydrolysis products do. This makes borohydride reduced cello-oligosaccharides valuable substrates for the assay of cellulase using traditional reducing sugar assays such as the Nelson-Symogyi method.
Dyed polysaccharide substrates
These substrates can be subdivided into two classes-
* Insoluble chromogenic substrates: An insoluble cellulase substrate such as AZCL-HE-cellulose absorbs water to create gelatinous particles when placed in solution. This substrate is gradually depolymerised and solubilised by the action of cellulase. The reaction is terminated by adding an alkaline solution to stop enzyme activity and the reaction slurry is filtered or centrifuged. The colour in the filtrate or supernatant is measured and can be related to enzyme activity.
* Soluble chromogenic substrates: A cellulase sample is incubated with a water-soluble substrate such as azo-CM-cellulose, the reaction is terminated and high molecular weight, partially hydrolysed fragments are precipitated from solution with an organic solvent such as ethanol or methoxyethanol. The suspension is mixed thoroughly, centrifuged, and the colour in the supernatant solution (due to small, soluble, dyed fragments) is measured. With the aid of a standard curve, the enzyme activity can be determined.
Enzyme coupled reagents
![Use of enzyme coupled reagents for the measurement of endo-cellulase](https://upload.wikimedia.org/wikipedia/commons/d/d8/Use_of_enzyme_coupled_reagents_for_the_measurement_of_endo-cellulase.svg)
Recently, new reagents have been developed that allow for the specific measurement of ''endo''-cellulase.
These methods involve the use of functionalised oligosaccharide substrates in the presence of an ancillary enzyme. In the example shown, a cellulase enzyme is able to recognise the trisaccharide fragment of cellulose and cleave this unit. The ancillary enzyme present in the reagent mixture (β-glucosidase) then acts to hydrolyse the fragment containing the chromophore or fluorophore. The assay is terminated by the addition of a basic solution that stops the enzymatic reaction and deprotonates the liberated phenolic compound to produce the phenolate species. The cellulase activity of a given sample is directly proportional to the quantity of phenolate liberated which can be measured using a spectrophotometer. The acetal functionalisation on the non-reducing end of the trisaccharide substrate prevents the action of the ancillary β-glucosidase on the parent substrate.
See also
*
Cellulose 1,4-beta-cellobiosidase
Cellulose 1,4-beta-cellobiosidase (, exo-cellobiohydrolase, beta-1,4-glucan cellobiohydrolase, beta-1,4-glucan cellobiosylhydrolase, 1,4-beta-glucan cellobiosidase, exoglucanase, avicelase, CBH 1, C1 cellulase, cellobiohydrolase I, cellobiohydrolas ...
, an efficient cellulase
*
Cellulase unit, a unit for quantifying cellulase activity
References
Further reading
*
* The Merck Manual of Diagnosis and Therapy, Chapter 24
*
*
{{Glycoside hydrolases
Enzymes
Cellulose