Subtilisin is a
protease
A protease (also called a peptidase, proteinase, or proteolytic enzyme) is an enzyme that catalyzes (increases reaction rate or "speeds up") proteolysis, breaking down proteins into smaller polypeptides or single amino acids, and spurring the form ...
(a
protein-digesting 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 ...
) initially obtained from ''
Bacillus subtilis
''Bacillus subtilis'', known also as the hay bacillus or grass bacillus, is a Gram-positive, catalase-positive bacterium, found in soil and the gastrointestinal tract of ruminants, humans and marine sponges. As a member of the genus '' Baci ...
''.
Subtilisins belong to
subtilases, a group of
serine proteases that – like all serine proteases – initiate the
nucleophilic attack on the
peptide (amide) bond through a serine
residue at the
active site. Subtilisins typically have molecular weights 27kDa. They can be obtained from certain types of
soil
Soil, also commonly referred to as earth or dirt, is a mixture of organic matter, minerals, gases, liquids, and organisms that together support life. Some scientific definitions distinguish ''dirt'' from ''soil'' by restricting the former ...
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 am ...
, for example, ''
Bacillus amyloliquefaciens'' from which they are secreted in large amounts.
Nomenclature
Subtilisin is also commercially known as ''Alcalase®'', ''Endocut-02L'', ''ALK-enzyme'', ''bacillopeptidase'', ''Bacillus subtilis alkaline proteinase bioprase'', ''bioprase AL'', ''colistinase'', ''genenase I'', ''Esperase®'', ''maxatase'', ''protease XXVII'', ''thermoase'', ''superase'', ''subtilisin DY'', ''subtilopeptidase'', ''SP 266'', ''Savinase®'', ''kazusase'', ''protease VIII'', ''protin A 3L'', ''Savinase®'', ''orientase 10B'', ''protease S.'' It is the type
serine endopeptidase of MEROPS family S8.
Structure
The structure of subtilisin has been determined by
X-ray crystallography
X-ray crystallography is the experimental science determining the atomic and molecular structure of a crystal, in which the crystalline structure causes a beam of incident X-rays to diffract into many specific directions. By measuring the angle ...
. The mature form is a 275-residue
globular protein
In biochemistry, globular proteins or spheroproteins are spherical ("globe-like") proteins and are one of the common protein types (the others being fibrous, disordered and membrane proteins). Globular proteins are somewhat water-soluble (fo ...
with several
alpha-helices, and a large
beta-sheet. The N-terminal contains an I9 propetide domain () that assists the folding of subtilisin. Proteolytic removal of the domain activates the enzyme. It is structurally unrelated to the
chymotrypsin-clan of serine proteases, but uses the same type of
catalytic triad in the
active site. This makes it a classic example of
convergent evolution.
Mechanism of catalysis
The active site features a charge-relay network involving Asp-32, His-64, and active site Ser-221 arranged in a
catalytic triad. The charge-relay network functions as follows: The carboxylate side-chain of Asp-32 hydrogen-bonds to a nitrogen-bonded proton on His-64's
imidazole ring. This is possible because Asp is negatively charged at physiological
pH. The other nitrogen on His-64 hydrogen-bonds to the O-H proton of Ser-221. This last interaction results in charge-separation of O-H, with the oxygen atom being more nucleophilic. This allows the oxygen atom of Ser-221 to attack incoming substrates (i.e., peptide bonds), assisted by a neighboring carboxyamide side-chain of Asn-155.
Even though Asp-32, His-64, and Ser-221 are sequentially far apart, they converge in the
3D structure to form the active site.
To summarize the interactions described above, Ser-221 acts as a
nucleophile and cleaves
peptide bond
In organic chemistry, a peptide bond is an amide type of covalent chemical bond linking two consecutive alpha-amino acids from C1 (carbon number one) of one alpha-amino acid and N2 ( nitrogen number two) of another, along a peptide or protein c ...
s with its partially negative oxygen atom. This is possible due to the nature of the charge-relay site of subtilisin.
Applications
Research tool
In molecular biology using ''B. subtilis'' as a
model organism, the gene encoding subtilisin (''aprE'') is often the second gene of choice after ''amyE'' for integrating reporter constructs into, due to its dispensability.
Commercial
Protein-engineered subtilisins are widely used in commercial products (the native enzyme is easily inactivated by detergents and high temperatures) and is also called a stain cutter, for example, in laundry and dishwashing
detergents,
cosmetics,
food processing
Food processing is the transformation of agricultural products into food, or of one form of food into other forms. Food processing includes many forms of processing foods, from grinding grain to make raw flour to home cooking to complex industr ...
, skin care products,
contact lens cleaners, and for research in
synthetic organic chemistry.
Occupational safety and health
People can be exposed to subtilisin in the workplace by breathing it in, swallowing it, skin contact, and eye contact. The
National Institute for Occupational Safety and Health
The National Institute for Occupational Safety and Health (NIOSH, ) is the United States federal agency responsible for conducting research and making recommendations for the prevention of work-related injury and illness. NIOSH is part of the ...
(NIOSH) has set a
recommended exposure limit (REL) of 60 ng/m
3 over a 60-minute period.
Subtilisin can cause "enzymatic detergent asthma". People who are sensitive to Subtilisin (Alcalase) usually are also allergic to the bacteria ''Bacillus subtilis''.
[ Mosby's Medical, Nursing, & Allied Health Dictionary, 14th edition, page 557 ]
References
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