Maize branching enzymes

The substrates, either 1 mg/ml amylose (c.l. 405) or amylopectin (c.l. 21), were incubated with 5 units/ml maize-branching enzyme (measured as by stimulation of phosphorylase a Guan and Preiss. 1993). The decrease in absorbance by the iodine/glucan complex was measured at 660 nm. 

V. HOW MANY GENES FOR THREE MAIZE-BRANCHING ENZYMES ... 

FIG. 5. Tryptic peptide maps of the maize-branching enzymes showing the similarity between the pair BEIIa/BEIIb and differences between the pair and BEI. Approximately 1.25 nmol of peptide digest was chromatographed on a reverse-phase column. Figure reprinted with permission from Singh and Preiss (1985). 

Baba, T., Kimura. K., Mizuno, K., Etoh, H., Ishida, Y., Shida. O., and Arai, Y. 1991. Sequence conversion of the catalytic regions of the amyiolytic enzymes in maize branching enzyme. Biochem. Biophys. Res. Commun. 181, 87-94. 

Cao, H and Preiss, J. 1996. Evidence for essential arginine residues at tre active site of maize branching enzymes. J. Protein Chem. 15, 291-304. 

Guan H. P., Kuriki, T., Sivak, M., and Preiss, J. 1995. Maize branching enzyme catalyzes synthesis of glycogen-like polysaccharide in glgB-deficient Escherichia coli. Proc. Natl. Acad. Science USA 92, 964-967. 

Potato branching enzyme thus shows a high degree of similarity to maize BEI and to maize BEII isozymes, however to a lesser extent (128, 129). As observed with the maize-branching enzymes, potato BEI was more active on amylose than BEII, and BEII was more active on amylopectin than BEI (130-132). 

Enzymes Involved in Starch Biosynthesis. Much of the eady data dealing with starch biosynthesis in plants are derived from the study of various mutants. The shrunken-2 and britde-2 mutants of maize have gready reduced levels of ADPGPP activity owing to the absence of one of the two subunits of this enzyme, and result in a shrunken seed appearance. Mendel s eady work on inheritance of traits was performed with a pea mutant deficient in branching enzyme activity (61). Mutations in plants affecting starch biosynthesis can have severe results to plant morphology and viability. 

Phytoglycogen-branching enzyme has been found in du however, no phytoglycogen was isolated by Black et al.23 Preiss and Boyer396 reported that the du mutation lowered starch synthase II activity and also lowered branching enzyme Ha activity. Gao et al.397 used a molecular approach to clone the du gene in maize endosperms and, based on amino acid sequence similarity of the predicted protein product with the soluble starch synthase III of potato,398 concluded that du most likely encodes the 180 000 molecular weight, primer-dependent soluble starch synthase described previously.399,400,401... 

The first enzymic studies done on the dul mutant were carried out in 1981, and both SSII and starch-branching enzyme Ha (SBEIIa) were found to have reduced activity in the endosperm compared to normal maize endosperm.204 SSII was shown to be different from SSI.173,205 SSII requires a primer for activity, and could not catalyze an unprimed reaction even in the presence of 0.5 M citrate, it also has less affinity for amylopectin than does SSI. However, 0.5 M citrate lowered the Km for amylopectin 17-fold. The activity with glycogen as a primer is one-half that observed with amylopectin. Therefore, glycogen is not as effective as a primer as is amylopectin, which differs from what was observed for starch synthase I. Both maize endosperm SSI and SSII had a Km for ADPGlc of 0.1 mM.196,205... 

Three forms of branching enzyme from developing hexaploid wheat (Triticum aestivum) endosperm have been partially purified and characterized.260 Two forms are immunologically related to maize BEI and one form to maize BEII. The N-terminal sequences are consistent with these relationships. The wheat BEIb gene is located on chromosome 7B, while the wheat BEIad peptide genes are located on chromosomes 7 A and 7D. The BE classes in wheat are differentially expressed during endosperm development. BEII is constitutively expressed throughout the whole cycle, while BEIb and BEIad are only expressed in late endosperm development. 

Amino acid sequence relationships between that of branching enzyme (BE) and amy-lolytic enzymes, such as a-amylase, pullulanase, glucosyltransferase and cyclodextrin glucanotransferase, especially at those amino acids believed to be contacts between the substrate and the amylase family enzymes, were first reported by Romeo et al.283 Baba et al.284 reported that there was a marked conservation in the amino acid sequence of the four catalytic regions of amylolytic enzymes in maize endosperm BEI. As shown in Table 4.12, four regions that putatively constitute the catalytic... 

As indicated in Table 4.12, four regions which constitute the catalytic regions of amylolytic enzymes are conserved in the starch-branching isoenzymes of maize endosperm, rice seed and potato tuber, and the glycogen-branching enzymes of E. coli.286,281 It would be of interest to know whether the seven highly conserved amino acid residues of the a-amylase family listed in bold letters in Table 4.12 are also functional in branching enzyme catalysis. Further experiments, such as chemical modification and analysis of the three-dimensional structure of the BEs, would be needed to determine the nature of its catalytic residues and mechanism. 

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