Barley purification

THE GLUTAMATE 1-SEMIALDEHYDE AMINOTRANSFERASE OF BARLEY PURIFICATION OF THE PROTEIN, c-DNA CLONING, SEQUENCING AND CHARACTERISATION OF THE mRNA... 

The Glutamate 1-Semialdehyde Aminotransferase of Barley Purification of the Protein, c-DNA Qoning, Sequencing and Characterisation of the mRNA 585... 

In 1833 an amylase from germinating barley was recovered and called diastase (1). Like malt itself, this product converted gelatinized starch into sugars, primarily maltose. Shordy thereafter, BerzeHus proclaimed the existence of non-living catalysts, and Schwaim (2) reported on his observation and purification of pepsin. 

K. Higuchi, K. Kanazawa, N. Nishizawa, M. Chino, and S. Mori, Purification and characterization of nicotianamine synthase from Fe-deficient barley roots. Plant Soil 765 173 (1994). 

Examination of the ratios of the dextrinogenic to the saccharogenic activities of malted barley extracts before and after treatment shows that the results of the Ohlsson procedures23 are not always predictable.8 The concentration of the amylases in the extracts, and the kinds and concentrations of substances which accompany them, influence the results. The presence or the absence of calcium ions is an important factor. Calcium ions increase the inactivation of beta amylase of malted barley and protect the alpha amylase from inactivation at unfavorable temperatures and also at unfavorable hydrogen ion activities.28 With purification, both amylases become increasingly thermolabile and increasingly sensitive to unfavorable hydrogen ion activities.78... 

Kristensen BK, Bloch H, Rasmussen SK (1999) Barley coleoptile peroxidases. Purification, molecular cloning, and induction by pathogens. Plant Physiol 120 501-512 Kubanek J, Jensen PR, Keifer PA, Sullards MC, Collins DO, Fenical W (2003) Seaweed resistance to microbial attack a targeted chemical defense against marine fungi. Proc Natl Acad Sci USA 100 6916-6921... 

T. B. Frandsen, F. Lok, E. Mirgorodskaya, P. Roepstorff, and B. Svensson, Purification, enzymatic characterisation and nucleotide sequence of a high-isoelectric-point a-glucosidase from barley malt, Plant Physiol., 123 (2000) 275-286. 

McMorrow, E.M. Bradbeer, J.W. Separation, purification, and comparative properties of chloroplast and cytoplasmic phosphoglycerate kinase from barley leaves. Plant Physiol., 93, 374-383 (1990)... 

Vasanthan, T. and Bhatty. R.S. 1995. Starch purification after pin milling and air classification of waxy, normal, and high amylose barleys. Cereal Chem. 72 379-384. 

Lee, R. C., Burton, R. A., Hrmova, M., and Fincher, G. B., Barley arabinoxylan arabinofuranohydrolases purification, characterization and determination of primary structures from cDNA clones. Biochem 72001, 356 (Pt 1), 181-9. 

Kotake, T., Nakagawa, N., Takeda, K., and Sakurai, N., Purification and characterization of wall-bound exo-1,3-beta-D-glucanase from barley (Hordeum vulgare L.) seedlings. Plant Cell Physiol 1997, 38 (2), 194-200. 

Additions of valine, leucine, and isoleucine were found to relieve the growth inhibitory effects of TP on Bacillus cell cultures, soybean cell cultures, and Arabidopsis seedlings. ALS isolated from a number of sources was found to be sensitive to TP at nM levels. The barley enzyme has been amenable to purification. A purification procedure that gives >60 % recovery and 235-fold purification is described. 

SA in Table II - nmoles/hr/mg protein and N.D - not detectable. Tobacco, soy cultures and barley seedlings were the best source of ALS, both in terms of specific activity and total units. The enzyme preparations from all sources were unstable in buffer solutions in spite of protective thiol agents. The inactivation of ALS in the crude extract of tobacco showed a distinct biphasic kinetics, implying the presence of at least two isozymes (unpublished observations). The presence of two ALS genes in tobacco (29) and at least three in microorganisms (18) has also been noted by other workers. ALS from barley was most amenable to purification. Table III gives a profile for the rapid purification of this enzyme with high recovery. 

The overall recovery of the enzyme was >60 % with a 235-fold purification. The final preparation, however, was not homogenous. Barley has two isoforms of ALS which can be separated on a phenyl agarose column immediately after the ammonium sulfate precipitation. One of the forms does not bind to the column and was too unstable to attempt purification. The details of purification in Table III pertain to the fraction with affinity for phenyl agarose. 

Work with a variety of plant systems has shown that multiple forms of SSS are present. Studies on barley endosperm, pea seeds, wheat endosperm, sorghum seeds, teosinte seeds, spinach leaf, maize endosperm, potato tuber, and rice seed extracts have indicated the presence of at least two major forms of SSS (reviewed in Preiss and Levi, 1980 Preiss, 1988 Preiss and Sivak, 1996), designated as types I and II. In maize leaf (Dang and Boyer, 1988) and castor bean endosperm (Goldner and Beevers, 1989), only one form of starch synthase was found, but since no extensive purification was attempted, the possibility remained that existing multiforms were not separated. Indeed, Downton and Hawker (1973) did find two forms of starch synthase in maize leaf, and thus the issue of the number of forms... 

There have been only a few kinetic studies on plant /3-D-manno-sidases, because of the apparently limited distribution of this enzyme in various species, and also because of the paucity of attempts at purification. The enzyme from malted barley has been purified 41-fold by fractionation with ammonium sulfate, and chromatography302 on Biogel P-100, DEAE-cellulose, and CM-cellulose. Some properties of /3-D-mannosidases from various tissues are summarized in Table II. 

Figure 2. Radiochromatograms for -metabolites of WPh-6E0 eluted from RP-HPLC columns with linear gradients (25 min) left, water-soluble NH -treated fraction from excised barley leaves ]50-100% CH3OH at 2.0 mL/min ( )] right, purification of peak C (8 mm cartridge, 25-100% CH CN at 2.5 mL/min).

Sun, C, Sathish, P, Ahlandberg, S, Dieber, A and Jansson, C. Identification of four starch-branching enzymes in barley endosperm partial purification of forms I, Ila and lib. 1997 New Phytol. 137 215-222. 

Limit Dextrins from Barley Starch. In one experiment, barley starch paste was treated with malt extract, in a second experiment with purified malt amylase (Table XVII). The purified enzyme has yielded limit dextrins with chain lengths greater than those given by the malt extract, possibly because certain carbohydrases capable of attacking the limit dextrins have been removed in the purification or because of a lower stability of the purified enzyme. 

Barley field experiments, description for brassin evaluation, 12 Barley leaf cell ultrastructure, protective effect of brassinosteroids under saline stress, 150,152/153,154/ Bean first-intemode bioassay, 321 Bean second-intemode bioassay activities of compounds, 65,68,69-7Of application, 321 brassin purification, 3-4 brassinosteroids, description, 109-110 development, 321 scheme, 63,65,66/... 

Hrmova M, Harvey AJ, Wang J, Shirley NJ, Jones GP, Stone BA, Hoj PB, Fincher GB (1996) Barley /3-D-Glucan Exohydrolase with /J-D-Glucosidase Activity. Purification, Characterization, and Determination of Primary Structure from a cDNA Clone. J Biol Chem 271 5277... 

The cultures of P. convolvulus were grown on moist barley grains at room temperature for a period of 28-30 days. Methanol extraction of the infected grains, followed by solvent partitioning of the metabolite mixture, led to the separation of the steroid compounds from metabolites 3, 4, 5 and 6 which dissolve readily in basic aqueous soivent. While the final purification of metabolites 1 and 2 was easily achieved by silica gel chromatography, the purification of the polar metabolites 3-6... 

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