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Observations on the Specificity and Nomenclature of Starch Debranching Enzymes

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Observations on the Specificity and Nomenclature of Starch Debranching Enzymes 83 (Oyo Toshitsu Kagaku (J. Appl. Glycosci.), Vol. 44, No. 1, p. 83-85 (1997) ) Mini Review Observations on the Specificity and Nomenclature of Starch Debranching Enzymes David J. MANNERS Department of Biological Sciences, Heriot- Watt University (Edinburgh EH14 4AS, Scotland, UK) Although the current literature contains references to four plant debranching enzymes-isoamylase , limit dextrinase, pullulanase and R-enzyme, the last three names are alternatives for the same enzyme , and only two plant debranching enzymes are, in fact, known. Moreover , the name pullulanase includes enzymes from both plant and microbial sources, even though these differ significantly in their ability to act on glycogen. To avoid any possible confusion or ambiguity, it is suggested that the second plant debranching enzyme should be known by the original name of limit dextrinase , rather than R-enzyme, and that the name pullulanase should be confined to enzymes of bacterial origin . The debranching enzymes, which specifically drolyses at varying rates, amylopectin, amylo- hydrolyse the (1•¨6)-ƒ¿-D-glucosidic linkages in pectin ƒÀ-limit dextrin, ƒ¿-dextrins from both amylopectin, glycogen and related dextrins, amylopectin and glycogen, and pullulan. It has continue to be the subject of intensive research little or no action on glycogen or phytoglycogen and industrial utilization. Unfortunately, some under conditions in which amylopectin is readily aspects of their specificity and enzyme nomen hydrolysed.3,4> The function of the enzyme is to clature may still cause confusion. For example, degrade a-dextrins during the in vivo break- the current literature contains references to down of starch, so that the logical name is ƒ¿- four plant debranching enzymes-isoamylase, dextrin 6-glucanohvdrolase or limit dextrinase. limit dextrinase, pullulanase and R-enzyme. The second plant debranching activity to be The general reader may not realise that there discovered, R-enzyme, acted on amylopectin are, in fact, only two major plant debranching and its 3-limit dextrin,5) but not on glycogen.6~ It enzymes, and that the last three names are also acted on a-dextrins from both amylopectin alternatives for the same enzyme. and glycogen, and therefore showed limit dex There is general agreement that the name trinase activity.6) Further work in the 1970s isoamylase' describes an enzyme, EC 3.2.1.68, established that limit dextrinase and R-enzyme which hydrolyses the inter-chain linkages in activities were due to the same enzyme,3,4) and glycogen, amylopectin and certain derived dex- the name limit dextrinase was preferred in trins, but has no action on pullulan. However, subsequent work since it related to the natural classification of the second plant debranching substrate.7-10) enzyme within the present Enzyme Nomencla In 1971, LEE and WHELAN11) suggested that ture Reports is difficult, and the descripition of there were only two classes of direct debranch EC 3.2.1.41 in both the 1992 and 1978 Reports is ing enzyme, isoamylase and pullulanase, and not entirely correct.1,2) included enzymes of both plant and microbial Limit dextrinase has been purified from many origin in the latter category. However, in con- higher plants, particularly cereals, and hy- trast to plant limit dextrinases, bacterial pul * Address for correspondence: 165 Mayfield Road , lulanases have a significant although incomplete Edinburgh EH9 3AY, Scotland, UK. action on glycogen. Whilst the inability of one 84 Oyo Toshitsu Kagaku (J. Appl. Glycosci.), Vol. 44, No. 1 (1997) a Within each of the three types of debranching enzyme , there are some variations in the rates of hydrolysis of particular substrates, depending on the exact source of the enzyme, but their activities are, in general, in accord with the above classification. Bacterial amylolytic enzymes which hydrolyse both (14) and (16) glucosidic linkages in certain substrates are excluded from this discussion. b An alternative but less preferred name is R-enzyme. C +, readily hydrolysed; •}, slowly hydrolysed; -, not hydrolysed. d Branched a-dextrins with side chains containing more than one glucose residue. Those a-dextrins with maltosyl side chains are more readily hydrolysed by limit dextrinase and pullulanase than by isoamylase. bacterial pullulanase preparation to act on par always be accurate. In other experiments,22) ticular samples of human liver glycogen12) and when debranching enzymes from potatoes and rabbit liver glycogen13) has been reported, the Aerobacter aerogenes which normally acted on same enzyme preparation attacked shellfish amylopectin, pullulan and a-dextrins were dilut glycogen to a significant extent.13) The general ed, the activity towards amylopectin selectively view from numerous publications is that pul disappeared. This was not due to a change in lulanases attack both amylopectin and gly enzyme specificity, but was due to the limit of cogen.l4-18) It is therefore suggested that the sensitivity of the analytical reagent at low rates name pullulanase should be restricted to of hydrolysis.23' The above two examples show enzymes of microbial origin which act, to vary- that specificity studies on debranching enzymes ing extents, on amylopectin, glycogen, pullulan are unusually sensitive to the exact experimen- and related dextrins. This would also have the tal conditions, as compared with the normal w- advantage that pullulanase, isopullulanase19) and j3-amylases. and neopullulanase20) would form a small sub- With the recognition that the debranching class of three enzymes, all of microbial origin, enzymes form part of the amylase 'super-fam which catalyse the depolymerization of pullulan ily' of enzymes, together with the normal in three different ways. amylases, branching enzymes and certain trans A revised classification of the debranching ferases,24) and with the increasing numbers of enzymes is shown in the Table 1. primary structures of debranching enzymes The above scheme represents a clarification which have been published25) or are being inves of that described in the Enzyme Nomenclature tigated, it is essential that there should be no Reports1,2) and by LEE and WHELAN,11) even dubeity about their specificity and nomencla though it may not apparently cover all exam ture. ples in the literature. For example, germinat ing rice endosperm contains a limit dextrinase REFERENCES which hydrolysed oyster glycogen and phyto glycogen, but at rates which were only 4% 1) Enzyme Nomenclature Report, I. U. B. M. B, p. 351, and 2% of that towards pullulan.21) However, 355 (1992). the actual enzyme concentrations used were 2) Enzyme Nomenclature Report, I. U. B, p. 282, 287 (1978). higher than those in other studies, so that 3) D. J. MANNERS: Biochem. Soc. Trans., 3, 49-53 assessment of relative rates of activity may not (1975). Starch Debranching Enzymes 85 4) D. J. MANNERS: in Biochemistry of Storage Carbo Acta, 293, 197-202 (1973). hydrates in Green Plants, P. M. DEY and R. A. DIxoN, 16) K. HATA, M. HATA, M. HATA and K. MATSUDA: eds., Academic Press, London, p. 149-203 (1985). Denpun Kagaku, 30, 95-101 (1983). 5) P. N. HOBSON,W. J. WHELANand S. PEAT : J. Chem. 17) A. MISAKI and M. YANO: Biryoeiyouso Kenkyu, 2, Soc., 1451-1459 (1951). 161-168 (1985). 6) S. PEAT, W. J. WHELAN, P. N. HOBSON and G. J. 18) Y. SAKANO: in Handbook of Amylases and Related THOMAS: J. Chem. Soc., 4440-4445 (1954). Enzymes, The Amylase Research Society of Japan, 7) A. W. MACGREGOR,L. J. MACRI,S. W. SCHROEDERand eds., Pergamon Press, Oxford, p. 131-139 (1988). S. L. BAZIN: Cereal Chem., 71, 610-617 (1994). 19) Y. SAKANO,M. HIGUCHIand T. KOBAYASHI: Arch. 8) M. KRISTENSEN,B. SVENSSONand J. LARSEN: Proc. Biochem. Biophys., 153, 180-187 (1972). 24th European Brewery Convention, Oslo, p.37-43 20) T. KURIKI,S. OKADAand T. IMANAKA: J. Bacteriol., (1993). 170, 1554-1559 (1988). 9) M. J. SISSONS, R. C. M. LANCE and D. H. B. 21) K. IWAKIand H. FUWA : Agric. Biol. Chem., 45, 2683- SPARROW: J. Cereal Sci., 17,19-24 (1993). 2688 (1981). 10) M. A. LONGSTAFFand J. H. BRYCE: Plant Physiol., 22) G. S. DRUMMOND,E. E. SMITH and W. J. WHELAN: 101, 881-889 (1993). FEBS Lett., 9, 136-140 (1970). 11) E. Y. C. LEE and W. J. WHELAN: in The Enzymes, 23) G. DUNN, D. G. HARDIEand D. J. MANNERS: Bio 3rd ed., Vol. 5, P. D. BoYER, ed., Academic Press, chem. J., 133, 413-416 (1973). New York, p. 191-234 (1971). 24) B. HENRISSAT: Biochem. J., 280, 309-316 (1991); H. 12) C. MERCIERand W. J. WHELAN: Eur. J. Biochem., 16, M. JESPERSEN,E. A. MACGREGOR,B. HENRISSAT,M. R. 579-583 (1970). SIERKSand B. SVENSSON: J. Protein Chem., 12, 791- 13) C. MERCIER,B. M. FRANTZand W. J. WHELAN: Eur. 805 (1993). J. Biochem., 26, 1-9 (1972). 25) For example, Y. NAKAMURA,T. UMEMOTO, N. 14) G. N. BATHGATEand D. J. MANNERS: Biochem. J., OGATA,Y. KUBOKI,M. YANGand T. SASAKI: Planta, 107, 443-445 (1968). 199, 209-218 (1996). 15) K. YOKOBAYASHI,H. AKAI, T. SUGIMOTO,M. HIRAO, K. SUGIMOTOand T. HARADA: Biochim. Biophys. (Received December 3, 1996).
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