Amylase: Difference between revisions
No edit summary |
Michal Harel (talk | contribs) No edit summary |
||
| (22 intermediate revisions by 4 users not shown) | |||
| Line 1: | Line 1: | ||
<StructureSection load=' | <StructureSection load='' size='350' side='right' scene='Sandbox_182/Alpha-amylase/1' caption='Amylase complex with Ca+2 (green) and Na+ (purple) ions (PDB code [[1hvx]])'> | ||
=Introduction= | =Introduction= | ||
Discovered and isolated by [http://en.wikipedia.org/wiki/Anselme_Payen Anselme Payen] in 1833, amylase was the first enzyme to be discovered<ref name="book">Yamamoto T.1988. Handbook of Amylases and Related Enzymes: Their Sources, Isolation Methods, Properties and Applications. Osaka Japan: Pergamon Press</ref>. Amylases are hydrolases, acting on α-1,4-glycosidic bonds<ref name="Path">PMID:9541387</ref>. They can be further subdivided into α,β and γ amylases<ref name="book"/>.'''α-Amylase''' (AAM) is an enzyme that acts as a catalyst for the hydrolysis of | Discovered and isolated by [http://en.wikipedia.org/wiki/Anselme_Payen Anselme Payen] in 1833, '''amylase''' was the first enzyme to be discovered<ref name="book">Yamamoto T.1988. Handbook of Amylases and Related Enzymes: Their Sources, Isolation Methods, Properties and Applications. Osaka Japan: Pergamon Press</ref>. Amylases are hydrolases, acting on α-1,4-glycosidic bonds<ref name="Path">PMID:9541387</ref>. They can be further subdivided into α,β and γ amylases<ref name="book"/>.'''α-Amylase''' (AAM) is an enzyme that acts as a catalyst for the hydrolysis of α-linked polysaccharides into α-anomeric products<ref name="Main">PMID:11226887</ref>. 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 saliva<ref name="Path"/>. α-Amylase is not only essential in human physiology but has a number of important biotechnological functions in various processing industries. '''β/α amylase''' (BAAM) is a precursor protein which is cleaved to form the β-amylase and α-amylase after secretion. '''β amylase''' (BAM) acts at the non-reducing chain ends and liberate only β-maltose<ref>PMID:6168260</ref>. '''γ amylase''' (GAM) acts at the non-reducing chain ends of amylose and amylopectin and liberates glucose. '''Pullulanase''' hydrolyses the α-1,6 glucoside linkage in starch, amylopectin, pullulan and related oligosaccharides<ref>PMID:22991654</ref>.<br /> | ||
*'''Neopullulanase''' is involved in starch degrading<ref>PMID:8955399</ref>.<br /> | |||
For α-amylase see [[Raghad zoubi]]<br /> | |||
See also [[Amylase (Hebrew)]]. | |||
=Structure<ref name="Main"/>= | =Structure<ref name="Main"/>= | ||
Shown as 1hvx is the structure of the thermostable α-amylase of ''Bacillus stearothermophilus'' (BSTA)<ref name="Main"/>. 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 <scene name='Sandbox_182/Domain_aa/1'>A domain </scene>constitutes the core structure, with a (β/α)<sub>8</sub>-barrel.The <scene name='Sandbox_182/Domain_a/1'> B domain</scene> 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 <scene name='Sandbox_182/Trio/1'>binding site</scene> for Ca<sup>2+</sup>-Na<sup>+</sup>-Ca<sup>2+</sup>. <scene name='Sandbox_182/Domain_c/1'>Domain C </scene>consisting of eight β-strands is assembled into a globular unit forming a Greek key motif. It also holds the <scene name='Sandbox_182/Caiii/1'>third </scene>Ca<sup>2+</sup> binding site in association with domain A. Positioned on the C-terminal side of the β-strands of the (β/α)<sub>8</sub>-barrel in domain A is the active site. The catalytic residues involved for the BSTA active site are | Shown as 1hvx is the structure of the thermostable α-amylase of ''Bacillus stearothermophilus'' (BSTA)<ref name="Main"/>. 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 <scene name='Sandbox_182/Domain_aa/1'>A domain </scene>constitutes the core structure, with a (β/α)<sub>8</sub>-barrel.The <scene name='Sandbox_182/Domain_a/1'> B domain</scene> 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 <scene name='Sandbox_182/Trio/1'>binding site</scene> for Ca<sup>2+</sup>-Na<sup>+</sup>-Ca<sup>2+</sup>. <scene name='Sandbox_182/Domain_c/1'>Domain C </scene>consisting of eight β-strands is assembled into a globular unit forming a Greek key motif. It also holds the <scene name='Sandbox_182/Caiii/1'>third </scene>Ca<sup>2+</sup> binding site in association with domain A. Positioned on the C-terminal side of the β-strands of the (β/α)<sub>8</sub>-barrel in domain A is the active site. The catalytic residues involved for the BSTA active site are <scene name='38/382954/Active_site_ball_stick/2'>Asp234, Glu264, and Asp331</scene>. The residues are identical to other α-amylases, yet there are positional differences which reflect the flexible nature of catalytic resides. | ||
<scene name=' | |||
<scene name='Sandbox_182/Trio/1'>CaII and CaI with Na</scene> found in the interior of domain B and <scene name='Sandbox_182/Caiii/2'>CaIII </scene>at the interface of domain A and C, constitute the metal ion binding sites. All α-amylases contain one strongly conserved Ca<sup>2+</sup> ion for structural integrity and enzymatic activity.<ref name="chloride">PMID: 12021442</ref> 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. | <scene name='Sandbox_182/Trio/1'>CaII and CaI with Na</scene> found in the interior of domain B and <scene name='Sandbox_182/Caiii/2'>CaIII </scene>at the interface of domain A and C, constitute the metal ion binding sites. All α-amylases contain one strongly conserved Ca<sup>2+</sup> ion for structural integrity and enzymatic activity.<ref name="chloride">PMID: 12021442</ref> 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== | ==Chloride Dependent Enzymes== | ||
| Line 23: | Line 25: | ||
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<ref name="book"/>. Dextrin, which is a viscosity improver, filler, or ingredient of food, is manufactured by the liquefaction of starch by bacteria α-amylase<ref name="book"/>. Bacterial α-amylases of ''B.subtilis'', or ''B.licheniformis'' are used for the initial starch liquefaction in producing high conversion glucose syrup<ref name="book"/>. 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<ref>PMID: 16286272 </ref>. α-Amylase is used for the production of malt, as the enzyme is produced during the germination of cereal grains<ref name="book"/>. | 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<ref name="book"/>. Dextrin, which is a viscosity improver, filler, or ingredient of food, is manufactured by the liquefaction of starch by bacteria α-amylase<ref name="book"/>. Bacterial α-amylases of ''B.subtilis'', or ''B.licheniformis'' are used for the initial starch liquefaction in producing high conversion glucose syrup<ref name="book"/>. 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<ref>PMID: 16286272 </ref>. α-Amylase is used for the production of malt, as the enzyme is produced during the germination of cereal grains<ref name="book"/>. | ||
β/α amylase (BAAM) is a precursor protein which is cleaved to form the β-amylase and α-amylase after secretion. | β/α amylase (BAAM) is a precursor protein which is cleaved to form the β-amylase and α-amylase after secretion. | ||
= | = Structure of the AmyC GH13 alpha-amylase from Alicyclobacillus sp, reveals accommodation of starch branching points in the alpha-amylase family<ref>doi 10.1107/S2059798318014900</ref> = | ||
The enzymatic degradation of starch has a myriad industrial applications. However, the branched nature of the polysaccharides that compose it poses problems, as branches have to be accommodated within an active centre best suited to linear polysaccharides. Alpha-amylases are glycoside hydrolases that break the α-1,4 bonds in starch and related glycans. The present work provides a rare insight into branch-point acceptance in these industrial catalysts. | |||
The complex of α-amylase from ''Alicyclobacillus sp.'' 18711 (AliC) with acarbose was solved by molecular replacement, with two molecules of AliC in the asymmetric unit, at a resolution of 2.1 Å ([[6gxv]]). The fold, as expected, is a canonical <scene name='79/799580/Cv1/5'>three-domain arrangement</scene> with the A, B and C domains defined approximately as <span style="color:deepskyblue;background-color:black;font-weight:bold;">A, residues 4–104 and 210–397 (in deepskyblue)</span>, <span style="color:yellow;background-color:black;font-weight:bold;">B, residues 105–209 (in yellow)</span>, and <span style="color:white;background-color:black;font-weight:bold;">C, residues 398–484 (in white)</span>. A classical Ca<sup>2+</sup>–Na<sup>+</sup>–Ca<sup>2+</sup> <scene name='79/799580/Cv1/4'>triad</scene> <ref name="Machius">PMID:9551551</ref>,<ref name="Brzozowski">PMID:10924103</ref> is found at the A/B-domain interface. The structure of AliC was determined in the presence of the <scene name='79/799580/Cv1/6'>inhibitor acarbose</scene> (<span style="color:lime;background-color:black;font-weight:bold;">colored in green</span>). As with many (retaining) α-amylase complexes, the acarbose is observed as a transglycosylated species, here a hexasaccharide which contains two of the acarviosin disaccharide motifs. The <scene name='79/799580/Cv/11'>complex defines six subsites</scene>, -4 to +2, with the expected catalytic GH13 signature triad of Asp234 (nucleophile), Glu265 (acid/base) and <scene name='79/799580/Cv/13'>Asp332 (interacting with O2/O3 of the -1 subsite sugar)</scene> all disposed for catalysis, here around the <sup>2</sup>H<sub>3</sub> half-chair of the unsaturated cyclohexitol moiety. AliC must also be able to accommodate branching in the +2 subsite, which is consistent with the <scene name='79/799580/Cv/14'>glucose moiety seen adjacent to O6 of the +2 sugar</scene>. | |||
*<scene name='79/799580/Cv/12'>Asp234 and Glu265 interactions</scene>. | |||
A ‘branched-ligand’ AliC complex was obtained through co-crystallization, with crystals forming in a new space group. This form diffracted poorly and data could only be obtained to 2.95 Å resolution [[6gya]]). Weak density in the -1 subsite, largely diffuse but greater than would be expected for discrete solvent, remained unmodelled. Density was clearer for a panose trisaccharide with an α-1,4-linked disaccharide in subsites +1 and +2 and, crucially, clear density for an α-1,6 branch accommodated in the +1 subsite, providing a structural context for the limit digest analysis of action on amylopectin starch. The <scene name='79/799580/Cv/15'>binding of the branched oligosaccharide in subsites +1, +2 and +1'</scene> (<span style="color:lime;background-color:black;font-weight:bold;">oligosaccharide colored in green</span>). | |||
=3D structures of amylase= | |||
[[Amylase 3D structures]] | |||
</StructureSection> | |||
=References= | =References= | ||
<references/> | <references/> | ||