Purification and characterization of soluble starch synthases from maize endosperm.
This study identified and characterized the soluble starch synthase of maize endosperm that was initially revealed as the SSII activity peak in anion exchange chromatography (J. L. Ozbun et al. (1971) Plant Physiol. 48, 765-769). At least six different genes coding for starch synthases are expressed in maize, although previously it was not known which of these is responsible for the SSII activity peak. The enzyme activity in the SSII peak was neutralized to a large extent by antibodies raised against the product of the Du1 gene, but was not affected by antibodies specific for the other highly expressed soluble starch synthase, zSSI, or for the zSSIIa or zSSIIb isoforms. These data provide direct evidence that Du1 codes for the starch synthase responsible for the SSII activity peak. This starch synthase was purified approximately 350-fold from endosperm extracts. The following enzymatic properties of the SSII activity were determined: temperature optimum, thermostability, pH effects, K(m) for different glucan primers and the glucosyl unit donor ADPGlc, V(max) using various primers, and stimulation by citrate. These properties were compared to those of zSSI purified over 1600-fold from maize endosperm by a parallel procedure. The major differences between the two enzymes were that the SSII activity displayed higher K(m) values for ADPGlc, a distinct temperature range for maximal activity, and different relative activities toward specific exogenous substrates. The purified SSI and SSII activities both were shown to be capable of elongating maltooligosaccharide primers in vitro.
Cao
H
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Department of Biochemistry, Iowa State University, 2110 Molecular Biology Building, Ames, Iowa, 50011, USA.
James
M G
MG
Myers
A M
AM
D004795:Enzyme Stability; D017343:Genes, Plant; D005964:Glucosyltransferases / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007120:Immunochemistry; D007527:Isoenzymes / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D007700:Kinetics; D009844:Oligosaccharides / Q000378:metabolism; D010940:Plant Proteins; D012995:Solubility; D013214:Starch Synthase; D013379:Substrate Specificity; D013696:Temperature; D003313:Zea mays / Q000201:enzymology* / Q000235:genetics
D007527:Isoenzymes; D009844:Oligosaccharides; D010940:Plant Proteins; C021705:maltooligosaccharides; D005964:Glucosyltransferases; C111377:starch synthase II; D013214:Starch Synthase