Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:2.4.1.18 (branching enzyme)
628 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Soluble starch synthase and branching enzyme were purified from 18-day-old cotyledons of the smooth-seeded pea cultivar Alaska (RR) and wrinkled-seeded pea cultivar Progress #9 (rr) by DEAE-cellulose chromatography. Two coeluting peaks of primed and citrate-stimulated starch synthase activity and a major and minor peak of branching enzyme activity were observed in Alaska. However, in Progress #9, only one peak of synthase activity was found. When crude extracts of Progress #9 were centrifuged, over 70% of the starch synthase activity was recovered in the pelleted fraction, and additional washings of the pellet released no further activity. The addition of purified starch granules to Alaska crude extracts also resulted in the recovery of a greater proportion of synthase activity in pelleted fractions. The two peaks of branching enzyme activity in Alaska differed in their stimulation of phosphorylase, amylose branching activity, and activity in various buffers. The DEAE-cellulose profile of Progress #9 showed no distinct peak of branching enzyme and less than 10% of the total activity found in Alaska. The association of one form of soluble starch synthase with the pelleted fraction and the greatly reduced levels of branching enzyme provide a partial explanation for the appearance of high-amylose starch in Progress #9 cotyledons.
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PMID:Soluble starch synthases and starch branching enzymes from cotyledons of smooth- and wrinkled-seeded lines of Pisum sativum L. 621 10

The structure of alpha-glucan, isolated from wild-type Escherichia coli B, a glycogen branching enzyme (BE)-deficient E. coli AC71 (glgB-), or from AC71 transformed with genes coding for maize BEI and BEII individually as well as with both genes, was analyzed by high-performance anion-exchange chromatography (HPAEC) with pulsed amperometric detection. Transformation of the maize BE gene(s) in AC71 (glgB-) showed complementation in branching activity. Analysis by HPAEC revealed different structures between glycogen of E. coli B and alpha-glucan of AC71 transformed with a different maize BE gene(s). The individual chains of the alpha-glucan debranched with isoamylase were distributed between chain length (CL) 3 and > 30 and the chain with CL 6 was the most abundant. In comparison with the glycogen of E. coli B, the alpha-glucan of AC71 transformed with the maize BE gene(s) consisted of a lesser amount of chains with CL 7-9 and a larger amount of chains with CL > 14. It also showed a broad peak with chains of CL 9-12 as in maize amylopectin. This study provides in vivo evidence that glycogen BE and maize BE isozymes may have different specificities in the length of chain transferred. Furthermore, this study suggests that the specificity of glycogen synthase and starch synthase and their concerted action with BE play an important role in determining the structure of the polysaccharide synthesized.
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PMID:Maize branching enzyme catalyzes synthesis of glycogen-like polysaccharide in glgB-deficient Escherichia coli. 786 74

Developing wild-type pea embryos contain two major isoforms of starch synthase and two isoforms of starch-branching enzyme. One of the starch synthases and both starch-branching enzymes occur both in the soluble fraction and tightly bound to starch granules. The other starch synthase, which is very similar to the waxy proteins of other species, is exclusively granule-bound., It is inactive when solubilized in a native form from starch granules, but activity is recovered when the SDS-denatured protein is reconstituted from polyacrylamide gels. Evidence is presented which indicates that all of these proteins become incorporated within the structure of the granule as it grows. It is proposed that the granule-bound waxy protein is active in vivo at the granule surface, whereas the remaining proteins are active in the soluble fraction of the amyloplast. The proteins become trapped within the granule matrix as the polymers they synthesize crystallize around them, and they probably play no further part in polymer synthesis.
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PMID:Soluble isoforms of starch synthase and starch-branching enzyme also occur within starch granules in developing pea embryos. 822 Apr 72

Antibodies were used to probe the degree of association of starch biosynthetic enzymes with starch granules isolated from maize (Zea mays) endosperm. Graded washings of the starch granule, followed by release of polypeptides by gelatinization in 2% sodium dodecyl sulfate, enables distinction between strongly and loosely adherent proteins. Mild aqueous washing of granules resulted in near-complete solubilization of ADP-glucose pyrophosphorylase, indicating that little, if any, ADP-glucose pyrophosphorylase is granule associated. In contrast, all of the waxy protein plus significant levels of starch synthase I and starch branching enzyme II (BEII) remained granule associated. Stringent washings using protease and detergent demonstrated that the waxy protein, more than 85% total endosperm starch synthase I protein, and more than 45% of BEII protein were strongly associated with starch granules. Rates of polypeptide accumulation within starch granules remained constant during endosperm development. Soluble and granule-derived forms of BEII yielded identical peptide maps and overlapping tryptic fragments closely aligned with deduced amino acid sequences from BEII cDNA clones. These observations provide direct evidence that BEII exits as both soluble and granule-associated entities. We conclude that each of the known starch biosynthetic enzymes in maize endosperm exhibits a differential propensity to associate with, or to become irreversibly entrapped within, the starch granule.
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PMID:Physical association of starch biosynthetic enzymes with starch granules of maize endosperm. Granule-associated forms of starch synthase I and starch branching enzyme II. 875 83

The aim of the work described in this paper was to characterize the tubers of potato (Solanum tuberosum var. Prairie) plants that had been transformed with the Escherichia coli ADPglucose pyrophosphorylase (EC 2.7.7.27) gene, glgC-16, under the control of a patatin promoter. Over 30 lines of transformed plants with increased ADPglucose pyrophosphorylase activity were obtained. The tubers of six of these lines were compared with those of control plants expressing the gene for beta-glucuronidase. The average increase in pyrophosphorylase activity was 200%, and the highest was 400%. Western immunoblotting of tuber extracts showed that the amounts of glgC-16 protein were linearly related to the extractable activity of the ADPglucose pyrophosphorylase. Cell fractionation studies showed that the increased activity of the pyrophosphorylase in the glgC-16 tubers had a similar intracellular location, the amyloplast fraction, to that found in the control tubers. No pleiotropic changes in the maximum catalytic activities of the following enzymes could be detected in the glgC-16 tubers: sucrose synthase, fructokinase, UDPglucose pyrophosphorylase, phosphofructokinase, soluble starch synthase, starch branching enzyme, phosphoglucomutase and alkaline inorganic pyrophosphatase. The glgC-16 tubers are held to be suitable for the study of the role of ADPglucose pyrophosphorylase in the control of starch synthesis.
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PMID:Characterization of transgenic potato (Solanum tuberosum) tubers with increased ADPglucose pyrophosphorylase. 897 57

A chromosomal region of Bacillus stearothermophilus TRBE14 which contains genes for glycogen synthesis was cloned and sequenced. This region includes five open reading frames (glgBCDAP). It has already been demonstrated that glgB encodes branching enzyme (EC 2.4.1.18 [H. Takata et al., Appl. Environ. Microbiol. 60:3096-3104, 1994]). The putative GlgC (387 amino acids [aa]) and GlgD (343 aa) proteins are homologous to bacterial ADP-glucose pyrophosphorylase (AGP [EC 2.7.7.27]): the sequences share 42 to 70% and 20 to 30% identities with AGP, respectively. Purification of GlgC and GlgD indicated that AGP is an alpha2beta2-type heterotetrameric enzyme consisting of these two proteins. AGP did not seem to be an allosteric enzyme, although the activities of most bacterial AGPs are known to be allosterically controlled. GlgC protein had AGP activity without GlgD protein, but its activity was lower than that of the heterotetrameric enzyme. The GlgA (485 aa) and GlgP (798 aa) proteins were shown to be glycogen synthase (EC 2.4.1.21) and glycogen phosphorylase (EC 2.4.1.1), respectively. We constructed plasmids harboring these five genes (glgBCDAP) and assayed glycogen production by a strain carrying each of the derivative plasmids on which the genes were mutated one by one. Glycogen metabolism in B. stearothermophilus is discussed on the basis of these results.
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PMID:Characterization of a gene cluster for glycogen biosynthesis and a heterotetrameric ADP-glucose pyrophosphorylase from Bacillus stearothermophilus. 924 54

The maize dull1 (du1) gene is a determinant of the structure of endosperm starch, and du1- mutations affect the activity of two enzymes involved in starch biosynthesis, starch synthase II (SSII) and starch branching enzyme IIa (SBEIIa). Six novel du1- mutations generated in Mutator-active plants were identified. A portion of the du1 locus was cloned by transposon tagging, and a nearly full-length Du1 cDNA sequence was determined. Du1 codes for a predicted 1674-residue protein, comprising one portion that is similar to SSIII of potato, as well as a large unique region. Du1 transcripts are present in the endosperm during the time of starch biosynthesis, but the mRNA was undetectable in leaf or root tissue. The predicted size of the Du1 gene product and its expression pattern are consistent with those of maize SSII. The Du1 gene product contains two repeated regions in its unique N terminus. One of these contains a sequence identical to a conserved segment of SBEs. We conclude that Du1 codes for a starch synthase, most likely SSII, and that secondary effects of du1- mutations, such as reduction of SBEIIa, result from the primary deficiency in this starch synthase.
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PMID:Characterization of dull1, a maize gene coding for a novel starch synthase. 950 Nov 6

In the developing endosperm of monocotyledonous plants, starch granules are synthesized and deposited within the amyloplast. A soluble stromal fraction was isolated from amyloplasts of immature maize (Zea mays L.) endosperm and analyzed for enzyme activities and polypeptide content. Specific activities of starch synthase and starch-branching enzyme (SBE), but not the cytosolic marker alcohol dehydrogenase, were strongly enhanced in soluble amyloplast stromal fractions relative to soluble extracts obtained from homogenized kernels or endosperms. Immunoblot analysis demonstrated that starch synthase I, SBEIIb, and sugary1, the putative starch-debranching enzyme, were each highly enriched in the amyloplast stroma, providing direct evidence for the localization of starch-biosynthetic enzymes within this compartment. Analysis of maize mutants shows the deficiency of the 85-kD SBEIIb polypeptide in the stroma of amylose extender cultivars and that the dull mutant lacks a >220-kD stromal polypeptide. The stromal fraction is distinguished by differential enrichment of a characteristic group of previously undocumented polypeptides. N-terminal sequence analysis revealed that an abundant 81-kD stromal polypeptide is a member of the Hsp70 family of stress-related proteins. Moreover, the 81-kD stromal polypeptide is strongly recognized by antibodies specific for an Hsp70 of the chloroplast stroma. These findings are discussed in light of implications for the correct folding and assembly of soluble, partially soluble, and granule-bound starch-biosynthetic enzymes during import into the amyloplast.
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PMID:Polypeptides of the maize amyloplast stroma. Stromal localization of starch-biosynthetic enzymes and identification of an 81-kilodalton amyloplast stromal heat-shock cognate. 953 63

Starch granules from maize (Zea mays) contain a characteristic group of polypeptides that are tightly associated with the starch matrix (C. Mu-Forster, R. Huang, J.R. Powers, R.W. Harriman, M. Knight, G.W. Singletary, P.L. Keeling, B.P. Wasserman [1996] Plant Physiol 111: 821-829). Zeins comprise about 50% of the granule-associated proteins, and in this study their spatial distribution within the starch granule was determined. Proteolysis of starch granules at subgelatinization temperatures using the thermophilic protease thermolysin led to selective removal of the zeins, whereas granule-associated proteins of 32 kD or above, including the waxy protein, starch synthase I, and starch-branching enzyme IIb, remained refractory to proteolysis. Granule-associated proteins from maize are therefore composed of two distinct classes, the surface-localized zeins of 10 to 27 kD and the granule-intrinsic proteins of 32 kD or higher. The origin of surface-localized delta-zein was probed by comparing delta-zein levels of starch granules obtained from homogenized whole endosperm with granules isolated from amyloplasts. Starch granules from amyloplasts contained markedly lower levels of delta-zein relative to granules prepared from whole endosperm, thus indicating that delta-zein adheres to granule surfaces after disruption of the amyloplast envelope. Cross-linking experiments show that the zeins are deposited on the granule surface as aggregates. In contrast, the granule-intrinsic proteins are prone to covalent modification, but do not form intermolecular cross-links. We conclude that individual granule intrinsic proteins exist as monomers and are not deposited in the form of multimeric clusters within the starch matrix.
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PMID:Surface localization of zein storage proteins in starch granules from maize endosperm. Proteolytic removal by thermolysin and in vitro cross-linking of granule-associated polypeptides. 953 75

The enzymatic reactions of bacterial glycogen and plant starch synthesis are similar and some of the properties of the biosynthetic enzymes are compared. Regulation occurs at the synthesis of ADPglucose and in almost all cases, ADPglucose pyrophosphorylase, is allosterically activated about 10- to over 40-fold by glycolytic intermediates and inhibited by AMP, ADP or Pi. The activator specificity of the ADPglucose pyrophosphorylase varies with respect to the source of enzyme and can be correlated to the major assimilation pathway occurring in the organism. For example, ADPglucose pyrophosphorylases from plants and other oxygenic photosynthetic organisms are activated by 3-phosphoglycerate. Organisms using glycolysis for carbon assimilation have ADPglucose pyrophosphorylases with fructose-1,6-bis-phosphate as the major activator. Chemical modification and site-directed mutagenesis studies that have determined the activator binding sites for some enzymes are described. The structural genes of Escherichia coli ADPglucose pyrophosphorylase allosteric mutants which no longer require activator for activity have been isolated. Transformation of plant systems with an allosteric bacterial mutant gene (but not with the wild-type gene) increases their starch content. Transformed potato tubers can have 25-60% more starch than the normal tuber indicating the importance of allosteric regulation of ADPglucose synthesis. The increase of a normal plant product by transformation of the plant with a gene encoding the rate-limiting enzyme in starch synthesis is an important biotechnological advance and suggests the possibilities of changing starch composition (extent of branching and chain sizes) via transformation with the starch synthase and branching enzyme genes.
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PMID:ADPglucose pyrophosphorylase: basic science and applications in biotechnology. 970 99


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