Debranching enzyme specificity

The major enzymes that have been used for studying structures of starch polymers and fragments from them are the endo-acting a-amylases, the exo-acting glucoamylase and (3-amylases, and the debranching enzymes, isoamylases and pullulanases. These enzymes have varied and diverse specificities that have been extensively studied (see previous sections). 

Lee, Y. M., Marshall, and Whelan, 1971. The substrate specificity of amylopectin-debranching enzymes from sweet com. Arch. Biochem. Biophys. 143, 365. 

Manners, D. J. 1997. Observations on the specificity and nomenclature of starch debranching enzymes. J. Appl. Glycosci. 44, 83-85. 

To explain the variation in the phosphate content of starches, it is postulated that some plants contain a specific phosphatase such an enzyme would have to possess unusual powers of penetration, since R-en-zyme, for example, cannot attack the interior of an amylopectin molecule. Moreover, a considerable proportion of the phosphorus would be situated in outer chains there is no evidence for this. (5) The formation of waxy-type starches is attributed to a low activity of the debranching enzyme caused by the presence of an inhibitor(s) to the debranching mechanism or by a deficiency of the debranching enzyme. Waxy-type starches should, therefore, contain a higher degree of branching than that actually observed (about 5%). 

The enzyme from rabbit muscle may be extracted and crystallized readily, but many recrystallizations may be necessary to ensure freedom from such contaminants as alpha-amylase, debranching enzyme, and branching enzyme. Purification may be facilitated by the use of column chromatography on 0-(2-diethylaminoethyl) cellulose. The enzyme exists in two forms (a and h) that differ in their requirement for adenosine 5 -monophosphate as cofactor. The b form is inactive in the absence of cofactor, and may be converted into the active (adenosine 5 -monophos-phate-independent) a form by phosphorylation of a specific serine residue under the action of (contaminant-free) phosphorylase kinase with adenosine 5 -triphosphate as the phosphate donor. 

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