Glycogen debranching enzyme: Difference between revisions

Revision as of 15:34, 27 March 2012

Revision as of 16:57, 2 June 2012

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== Function ==

Together with [[phosphorylase]], glycogen debranching enzymes function in [[glycogenolysis|glycogen breakdown]] and glucose mobilization. When phosphorylase has digested a glycogen branch down to four glucose residues, it will not remove further residues. Glycogen debranching enzymes assist [[phosphorylase]], the primary enzyme involved in [[glycogen breakdown]], mobilize glycogen stores. Phosphorylase can only cleave α-1,4- glycosidic bond between adjacent glucose molecules in glycogen but branches exist as α-1,6 linkages. When phosphorylase reaches four residues from a branching point it stops cleaving; because 1 in 10 residues is branched cleavage by phosphorylase alone would not be sufficient in mobilizing glycogen stores. <ref name=Berg/><ref name=Hondoh/> Before phosphorylase can resume catabolism, debranching enzymes perform two functions:

* 4-α-D-glucanotransferase ({{EC number|2.4.1.25}}), or [[glucosyltransferase]], transfers three glucose [[residue (chemistry)|residues]] from the four-residue glycogen branch to a nearby branch. This exposes a single glucose residue joined to the glucose chain though an α -1,6 glycosidic linkage<ref name="Berg"/>

* Amylo-α-1,6-glucosidase ({{EC number|3.2.1.33}}), or [[glucosidase]], cleaves the remaining alpha-1,6 linkage, producing glucose and a linear chain of glycogen.<ref name=Berg/> The mechanism by which the glucosidase cleaves the α -1,6-linkage is not fully known because the [[amino acids]] in the [[active site]] have not yet been identified. It is thought to proceed through a two step acid base assistance type mechanism, with a [[oxocarbenium]] ion intermediate, and retention of configuration in glucose. <ref name=Molecule/> This is a common method through which to cleave bonds, with an acid below the site of [[hydrolysis]] to lend a proton and a base above to deprotinate a water which can then act as a [[nucleophile]]. These acids and bases are amino acid side chains in the active site of the enzyme. A scheme for the mechanism is shown in the figure below.<ref name=MCCarter/>

[[File:Glycosidase mechanism.png]]

Thus the debranching enzymes, transferase and α-1,6- glucosidase converts the branched glycogen structure into a linear one, paving the way for further cleavage by phosphorylase.

Thus the debranching enzymes, transferase and α-1,6- glucosidase converts the branched glycogen structure into a linear one, paving the way for further cleavage by phosphorylase.

== Structure and activity ==

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=== One enzyme with two catalytic sites ===

In mammals and [[yeast]], a single enzyme performs both debranching functions.<ref name="Nakayama"/> The human glycogen debranching enzyme (gene: AGL) is a monomer with a molecular weight of 175 kDa. It has been shown that the two catalytic actions of AGL can function independently of each other, demonstrating that multiple active sites are present. This idea has been reinforced with inhibitors of the active site, such as polyhydroxyamine, which were found to inhibit glucosidase activity while transferase activity was not measurably changed.<ref name=Gillard/> Glycogen debranching enzyme is the only known eukaryotic enzyme that contains multiple catalytic sites and is active as a monomer.<ref name="Chen"/><ref name="UniProt P35573"/>

Some studies have shown that the C-terminal half of yeast GDE is associated with glucosidase activity, while the N-terminal half is associated with glucosyltransferase activity.<ref name=Nakayama/> In addition to these two [[active site]]s, AGL appears to contain a third active site that allows it to bind to a glycogen polymer.<ref name="Gillard"/> Though the complete structure of the GDE in Eukaryotes is yet to be determined it is thought to bind to six glucose molecules of the chain as well as the branched glucose, thus corresponding to 7 subunits within the active site, as shown in the figure below.<ref name = Yamamoto/>

[[File:Hypothesized substraight binding ___location.png]]

It was seen that when glucose ‘a’, ‘b’, ‘c’ and ‘0’ in the [[active site]] was hydrolyzed the most rapidly.<ref name=Yamamoto/> This indicated that this region of the glycogen chain bond strongest to the active site because a stronger interaction between enzyme and substrate leads to a more rapid hydrolysis.

Despite these advances, the complete structure of GDE in eukaryotes has yet to be determined.<ref name="Woo"/> The glycogen-degrading enzyme of the [[archaea]] ''[[Sulfolobus solfataricus]]'' is better characterized than those of [[eukaryotes]]. The GDE of ''S. solfataricus'' is known as treX. Although, like mammalian GDE, treX has both amylosidase and glucanotransferase functions, TreX is structurally similar to glgX, and hass a mass of 80kD and one active site.<ref name=Woo/><ref name ="UniProt A8QX06"/> Unlike either glgX or AGL, however, treX exists as a dimer and tetramer in solution. TreX's oligomeric form seems to play a significant role in altering both enzyme shape and function. Dimerization is thought to stabilize a "flexible loop" located close to the active site. This may be key to explaining why treX (and not glgX) shows glucosyltransferase activity. As a tetramer, the catalytic efficiency of treX is increased fourfold over its dimeric form.<ref name=Song/><ref name="Park"/>

== Genetic Location ==

The official name for the gene is “amylo- α- 1,6- glucosidase, 4- α- glucanotransferase,” with the official symbol AGL. AGL is an autosomal gene found on chromosome lp21.<ref name=Hondoh/> The AGL gene provides instructions for making several different versions, known as isoforms, of the glycogen debranching enzyme. These isoforms vary by size and are expressed in different tissues, such as liver and muscle. This gene has been studied in great detail, because mutation at this gene is the cause of Glycogen Storage Disease Type III. <ref name=Gene/>

The gene is 85 kb long, has 35 [[exon]]s and encodes for a 7.0 kb- mRNA. Translation of the gene begins at exon 3,which encodes for the first 27 amino acids of the AGL gene, because the first two exons (68kb) contain the 5’ untranslated region. Exons 4-35 encode the remaining 1505 amino acids of the AGL gene. <ref name= Bao/>

Studies produced by the department of pediactrics at Duke University suggest that the human AGL gene contains at minimum 2 promotor regions, sites where the transcription of the gene begins, that result in differential expression of isoform, different forms of the same protein, mRNAs in a manner that is specific for different tissues.<ref name =Ding Gillard /><ref name = GillardDing />

== Pathology ==

{{Main|Glycogen storage disease type III}}

When GDE activity is compromised, the body cannot effectively release stored glycogen, type III Glycogen Storage Disease (debrancher deficiency), an autosomal recessive disorder, can result. In GSD III glycogen breakdown is incomplete and there is accumulation of abnormal glycogen with short outer branches. <ref name= Monga/>

Most patients exhibit GDE defiency in both liver and muscle (TypeIIIa), although 15% of patients have retained GDE in muscle while having it absent from the liver (Type IIIb).<ref name=Hondoh/> Depending on [[mutation]] ___location, different mutations in the AGL gene can effect different isoforms of the [[gene expression]]. For example, mutations that occur on exon 3, effect the form which affect the [[isoform]] that is primarily expressed in the liver; this would lead to GSD type III.<ref name=Shen/>

These different manifestation produce varied symptoms, which can be nearly indistinguishable from Type I GSD, including [[hepatomegaly]], [[hypoglycemia]] in children, short stature, [[myopathy]], and [[cardiomyopathy]].<ref name=Bao/><ref name=Talente/> Type IIIa patients often exhibit symptoms related to liver disease and progressive muscle involvement, with varationsvariations caused by age of onset, rate of disease progression and severity. Patients with Type IIIb generally symptoms related to liver disease.<ref name =Kishnan/> Type III patients be distinguished by elevated liver enzymes, with normal [[uric acid]] and blood lactate levels, differing from other forms of GSD.<ref name=Shen/> In patients with muscle involvement, Type IIIa, the muscle weakness becomes predominant into adulthood and can lead to ventricular [[hypertrophy]] and distal muscle wasting.<ref name=Shen/>

== References ==

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{{Glycosyltransferases}}

{{Sugar hydrolases}}

{{DEFAULTSORT:Glycogen Debranching Enzyme}}