Amylase

Introduction Discovered and isolated by Anselme Payen in 1833, amylase was the first enzyme to be discovered. Amylases are hydrolases, acting on α-1,4-glycosidic bonds. They can be further subdivided into α,β and γ amylases.α-Amylase (AAM) is an enzyme that acts as a catalyst for the hydrolysis of alpha-linked polysaccharides into α-anomeric products. The enzyme can be derived from a variety of sources, each with different characteristics. α-Amylase found within the human body serves as the enzyme active in pancreatic juice and salvia. α-Amylase is not only essential in human physiology but has a number of important biotechnological functions in various processing industries. Beta/alpha amylase (BAAM) is a precursor protein which is cleaved to form the beta-amylase and alpha-amylase after secretion. Structure Shown as 1hvx is the structure of the thermostable α-amylase of Bacillus stearothermophilus (BSTA). BSTA is comprised of a single polypeptide chain. This chain is folded into three domains: A, B and C. These domains are generally found on all α-amylase enzymes. The constitutes the core structure, with a (β/α)8-barrel.The consists of a sheet of four anti-parallel β-strands with a pair of anti-parallel β-strands. Long loops are observed between the β-strands. Located within the B domain is the for Ca2+-Na+-Ca2+. consisting of eight β-strands is assembled into a globular unit forming a Greek key motif. It also holds the Ca2+ binding site in association with domain A. Positioned on the C-terminal side of the β-strands of the (β/α)8-barrel in domain A is the active site. The catalytic residues involved for the BSTA active site are . The residues are identical to other α-amylases, yet there are positional differences which reflect the flexible nature of catalytic resides. found in the interior of domain B and at the interface of domain A and C, constitute the metal ion binding sites. All α-amylases contain one strongly conserved Ca2+ ion for structural integrity and enzymatic activity. CaI is consistent in α-amylases, however there are structural differences between the linear trio of CaI, CaII and Na in other enzymes. CaIII acts as a bridge between two loops, one from α6 of domain A, and one between β1 and β2 of domain C. Chloride Dependent Enzymes A family of chloride-dependent enzymes, including salivary and pancreatic α-amylase, require the binding of a chloride ion to be allosterically activated. The function of the chloride ion still remains uncertain. No relationship has been observed between the anion binding affinity and its activity, indicating the complexity between the binding parameters and mechanism it activates. Studies have shown that nitrite and nitrate ions with pancreatic α-amylase fit within the chloride binding site, thus making all the necessary hydrogen bonds and enhancing the relative activity by 5-fold. Function Mechanism In the human body, α-amylase is part of digestion with the breakdown of carbohydrates in the diet. The mechanism involved includes catalyzing substrate hydrolysis by a double replacement mechanism, forming a covalent glycosyl-enzyme intermediate and hydrolyzed through oxocarbenium ion-like transition states. One of the carboxylic acids in the active site acts as the catalytic nucleophile during the formation of the intermediate. A second carboxylic acid operates as the acid/base catalyst, supporting the stabilization of the transition states during the hydrolysis. Human Salivary and Pancreatic α-Amylase Salivary α-Amylase hydrolyzes the (α1-4) glycosidic linkages of starch, separating it into short polysaccharide fragments. Once the enzyme reaches the stomach, it becomes inactivated due to the acidic pH. Further breakdown of starch occurs by secretion of a second form of the enzyme by the pancreas. Pancreatic juice enters the duodenum and pancreatic α-amylase further cleaves starch to yield maltose, maltotriose and oligosaccharides. The oligosaccharides are referred to as dextrins, which are fragments of amylopectin consisting of (α1-6)branch points. Microvilli of the intestinal epithelia break maltose and dextrins into glucose, which gets absorbed into the circulatory system. Glycogen has a relatively similar structure as starch, and thus proceeds in the same digestive pathway. Regulation α-Amylase is regulated through a number of inhibitors. These inhibitors are classified according to six categories, based on their tertiary structures. Inhibitors of α-amylase block the active site of the enzyme. In animals, inhibitors control the conversion of starch to simple sugars during glucose peaks after a meal so that breakdown of glucose occurs at a rate the body can handle. This is particularly important for diabetics, who require low quantities of α-amylase to maintain control over glucose levels. After taking insulin however, pancreatic α-amylase escalates. Plants use these inhibitors as a defense mechanism to inhibit the use of α-amylase in insects, thus protecting themselves from herbivory. Industrial Uses α-Amylase is used extensively in various industrial processes. In textile weaving, starch is added for warping. After weaving, the starch is removed by Bacillus subtilis α-amylase. Dextrin, which is a viscosity improver, filler, or ingredient of food, is manufactured by the liquefaction of starch by bacteria α-amylase. Bacterial α-amylases of B.subtilis, or B.licheniformis are used for the initial starch liquefaction in producing high conversion glucose syrup. Pancreatitis can be tested by determining the level of amylases in the blood, a result of damaged amylase-producing cells, or excretion due to renal failure. α-Amylase is used for the production of malt, as the enzyme is produced during the germination of cereal grains. β/α amylase (BAAM) is a precursor protein which is cleaved to form the β-amylase and α-amylase after secretion.

spinqualitylabelsall models Amylase complex with Ca+2 (green) and Na+ (purple) ions (PDB code 1hvx) Show:Asymmetric Unit Biological Assembly Drag the structure with the mouse to rotate   Export Animated Image

3D structures of Amylase3D structures of Amylase

Updated on 24-August-2014

α-amylaseα-amylase

3ij7, 1xv8, 1c8q, 1bsi, 1smd, 1hny – hAAM – human

1xgz, 1q4n, 1kb3, 1kbb, 1kbk, 1kgu, 1kgw, 1kgx, 1jxj, 1jxk, 2cpu - hAAM (mutant)

3n8t – TkAAM – Thermococcus kodakarensis

3k8k – BtAAM – Bacterioides thetaiotaomicron

3kwx, 2guy – AoAAM – Aspergilluys oryzae

2wpg – AAM – Xanthomonas campestris

2wc7, 2wcs, 2wkg – AAM catalytic region – Cyanobacterium

3bh4 – BaAAM – Bacillus amyloliquefaciens

1e3x, 1e43 – BaAAM chimera

1ht6, 1amy - bAAM – barley

3bsg – bAAM (mutant)

3dhu – AAM – Lactobacillus plantarum

3dc0 – AAM – Bacillus KR8104

3bcf, 1wza – HoAAM – Halothermothrix orenii

2die, 1wp6 – AAM alkaline – Bacillus sp.

1ud2, 1ud4, 1ud5, 1ud6, 1ud8 - AAM – Bacillus sp. KSM-K38

1ud3 – AAM (mutant) – Bacillus sp. KSM-K38

2gjr – BhAAM – Bacillus halmapalus

2b5d – AAM – Thermotoga maritima

2c3g, 2c3v – BhaloAAM – Bacillus halodurans

1ji1, 1ji2, 1bvz - TvAAM – Thermoactinomyces vulgaris

1wzk, 1wzl, 1wzm, 1izj, 1izk, 1jf5, 1jf6 – TvAAM (mutant)

1mwo, 1mxd – PwAAM – Pyrococcus woesei

1ob0, 1bli - BlAAM (mutant) – Bacillus licheniformis
1vjs – BlAAM precursor
1bpl – BlAAM
1b0i, 1aqm, 1aqh – PhAAM - Pseudoalteromonas haloplanktis

1jd7, 1jd9 - PhAAM (mutant)

1g5a – NpAAM – Neisseria polysaccharea

1hvx - BaAAM – Bacillus stearothermophilus

1qho – GsAAM – Geobacillus stearothermophilus

1jae – TmAAM – Tenebrio molitor

1pif – pAAM – pig

6taa, 2aaa - AoAAM
4aee – AAM catalytic domain – Staphylothermus marinus
3ren – AAM – Clostridium perfringens
3vm5 – AAM – Oryzias latipes
3vm7 – AAM – Malbranchea cinnamomea
4gkl – AAM – Thermotoga neapolitana
4ays – AAM – Deinococcus radiodurans

AAM binary complexes

1xd0, 1xd1, 1cpu, 1jfh - hAAM + saccharide
1b2y, 1xcw, 1xcx - hAAM + acarbose
3blk, 3blp, 1z32, 1nm9, 1mfu, 1mfv, 3cpu - hAAM (mutant) + saccharide
3dhp, 1xh0, 1xh2 - hAAM (mutant) + acarbose
3ij8, 3ij9 – hAAM catalytic intermediate

2qmk, 3bai – hAAM + NO2

3baw – hAAM + N3

3bax - hAAM (mutant) + N3

3bak – hAAM (mutant) + NO3

1xh1 - hAAM (mutant) + Cl

2qv4, 3baj, 3bay - hAAM + acarbose + NO2

3old, 3ole, 3olg, 3oli – hAAM + statin

1u2y, 1u30, 1u33 – hAAM + inhibitor
4gqq – hAAM + ethyl caffeate
4gqr – hAAM + myricetin
3n92, 3n98 – TkAAM + saccharide
3l2l, 3l2m, 1vah, 1wo2, 1ua3, 1pig, 1ppi - pAAM + saccharide
1hx0 – pAAM + acarbose
1kxq, 1kxt, 1kxv – pAAM + antibody VHH fragment

1bvn, 1dhk – pAAM + protein inhibitor

1ose – pAAM + acarbose
3k8l - BtAAM (mutant) + saccharide
3k8m - BtAAM + acarbose
2d2o, 1jl8, 1jib - TvAAM + saccharide
3a6o – TvAAM + acarbose
2d0f, 2d0g, 2d0h, 1vb9, 1vfm, 1vfo, 1vfu, 1uh2, 1uh4 - TvAAM (mutant) + saccharide
1uh3 - TvAAM (mutant) + acarbose
1ava – bAAM + protein inhibitor

1bg9, 1rpk - bAAM + acarbose
1p6w – bAAM + substrate analog

1rp8, 1b1y, 1rp9 - bAAM (mutant) + saccharide
3bsh, 2qpu, 2qps - bAAM (mutant) + acarbose
3bcd - HoAAM + saccharide
3bc9 - HoAAM + acarbose
2gjp, 1w9x - BhAAM + saccharide
2gvy - AoAAM + saccharide
1zs2, 1s46, 1mvy, 1mw0, 1mw1, 1mw2, 1mw3 - NpAAM (mutant) + saccharide
1bag - BsAAM + saccharide – Bacillus subtilis

1ua7 – BsAAM + acarbose
2d3l, 2d3n - BacAAM + saccharide – Bacillus

2c3h, 2c3w, 2c3x - BhaloAAM + saccharide
1mxg - PwAAM + acarbose
3qgv - PwAAM + sucrose
1g9h, 1g94 - PhAAM + saccharide
1kxh - PhAAM (mutant) + acarbose
1l0p – PhAAM + NO3

1e40 – BaAAM chimera + saccharide
1e3z - BaAAM chimera + acarbose
1fa2 - AAM + saccharide – Sweet potato

1qhp - GsAAM + saccharide
4e2o - GsAAM + acarbose
1clv, 1viw, 1tmq – TmAAM + protein inhibitor

1gah, 1gai – AaAAM + acarbose – Aspergillus awamori

3gly, 1agm, 1glm – AaAAM + saccharide

Pullulanase α-amylasePullulanase α-amylase

2wan – AAM – Bacillus acidopullululyticus

2fgz – KaAAM – Klebsiella aerogenes

2e8y - BsAAM
1ji2 - TvAAM
1jl5, 1jf6, 1wzk, 1wzl, 1wzm - TvAAM (mutant)

Pullulanase α-amylase binary complexes

2fh6, 2fh8, 2fhb, 2fhc, 2fhf - KaAAM + saccharide
3fax - AAM + saccharide – Streptococcus agalactiae

2e8z, 2e9b - BsAAM + saccharide
2d2o - TvAAM + saccharide
1g1y, 1jib, 1jl8, 1vfm, 1vfo, 1vfu, 1vb9 - TvAAM (mutant) + saccharide
3a6o – TvAAM + acarbose

Neopullulanase α-amylaseNeopullulanase α-amylase

4aef – AAM – Pyrococcus furiosus
1j0h – BsAAM
2z1k – AAM – Thermus thermophilus
1j0i – BsAAM + α-D-glucose
1j0k – BsAAM (mutant) + α-D-glucose
1j0j – BsAAM (mutant) + maltotetraose

β-amylaseβ-amylase

2xfr – bBAM

1wdp – sBAM – soybean

2dqx, 1uko, 1ukp – sBAM (mutant)

1vem, 5bca, 1cqy, 1b90 – BcBAM – Bacillus cereus

1ven - BcBAM (mutant)
1fa2 - AAM + saccharide – Sweet potato

β-amylase binary complexes

2xff – bBAM + acarbose
2xfy, 2xg9, 2xgb, 2xgi – bBAM + inhibitor

1wdq, 1wdr, 1wds, 1v3h, 1v3i, 1q6d, 1q6e, 1q6f, 1q6g - sBAM (mutant) + saccharide
1q6c, 1bfn, 1bya, 1byb, 1byc, 1byd, 1btc - sBAM + saccharide
1j0y, 1j0z, 1j10, 1j11, 1j12, 1j18, 1b9z - BcBAM + saccharide
1veo, 1vep, 1itc - BcBAM (mutant) + saccharide
1b1y - AAM (mutant) + saccharide – Hordeum vulgare

γ-amylaseγ-amylase

1lf6 – TtGAM – Thermoanaerobacterium thermosaccharolyticum

1lf9 - TtGAM + acarbose

β/α-amylaseβ/α-amylase

2laa, 2lab – PpBAAM – Paenibacillus polymyxa - NMR
3voc – PpBAAM

Maltohexaose-producing amylaseMaltohexaose-producing amylase

1wp6 - BacMAM 1wpc – BacMAM + saccharide

Maltogenic amylaseMaltogenic amylase

1gvi, 1sma – MAM – Thermus sp.

Taka amylaseTaka amylase

2taa, 3vx0, 3vx1 – AoTAM

7taa – AoTAM + acarbose