PURIFICATION AND PROPERTIES OF A PHYTASE FROM RYE

Phytase (myo-inositol hexakisphosphate phosphohydrolase) has been purified about 2,000-fold from ungerminated rye with a recovery of 6%. The enzyme behaves as a monomeric protein of a molecular mass of about 67 kDa. OptimalpH for the degradation of phytate has been found at pH 6.0 and 45C. Kinetic parameters for the hydrolysis of Na-phytate are KM300 μM and kcat 358 s^?1 at 35C and pH 6.0. The rye enzyme exhibits a broad affinity for various phosphorylated compounds and hydrolyses phytate in a stepwise manner; the pentakis- and tetrakisphosphate were identified as 1(1,2,3,4,5)P₅ and I(2,3,4,5)P4 Consequently, this enzyme is a 6-phytase (EC 3.1.3.26).     

Response of Shiraz grapevines to five different training systems in the Barossa Valley, Australia

Own‐rooted Shiraz, spaced 1.5 m apart in 2.75 m wide rows, were compared under five different training systems in the Barossa Valley, South Australia over five seasons starting 1995–96. Our objective was to determine how training affected fruit composition and crop yield in a context of efficient vineyard management. Training systems were: (i) low single wire (LSW), in which vines were trained to bi‐lateral cordons, 1.0 m above the ground, non‐shoot‐positioned; (ii) high single wire (HSW), a higher cordon (1.8 m) version of LSW; (iii) vertically shoot positioned (VSP); (iv) Scott Henry, where canopies of alternate vines were cordon‐trained and shoot‐positioned upwards (SHU) or downwards (SHD); and (v) minimally pruned (MIN), the same as LSW, except vines were not annually spur‐pruned. Crop yields (kg/m of row), over four seasons, averaged 4.9 (MIN), 2.9 (combined Scott Henry), 3.2 (VSP), 2.6 (HSW) and 2.9 (LSW). The greatest year‐to‐year variation in yield occurred with MIN and Scott Henry training. Crop weight of SHD vines was inferior to that of SHU vines in two seasons due to fewer shoots and bunches per vine, and to fewer bunches per shoot. Individual berry weights (g) were consistently least with MIN (0.89), and greatest with VSP (1.17). Fruit from all training systems exhibited similar rates of sugar accumulation during two seasons in which repeated measures of fruit maturation were made. Excepting the relatively light yields observed in 1999–2000, sugar accumulation was delayed in MIN vines, relative to other training systems, even when MIN harvest was delayed up to two weeks. The delay is most readily explained by the greater crops of MIN vines. Berry total anthocyanins and total phenolics concentrations (mg/g berry fresh wt.) at harvest were not greatly affected by training system. Berry anthocyanins and total phenolics exhibited a negative relationship with crop/m of canopy and a slight positive relationship with bunch exposure when evaluated across all training systems. Experimental plot soil depth and water availability affected cane pruning weights, yield per vine, berry weight, and canopy characteristics. LSW, HSW and MIN training systems all provided good yields of high quality fruit, although MIN did have a tendency to produce excessive crops in some years. VSP and Scott Henry training were less attractive due to their inherently greater canopy management requirements.     

IMPROVING ENZYMATIC REDUCTION of MYO-INOSITOL PHOSPHATES WITH INHIBITORY EFFECTS ON MINERAL ABSORPTION IN BLACK BEANS (PHASEOLUS VULGARIS VAR. PRETO)

Activation of endogenous or addition of exogenous phytases during food processing gives a chance to reduce the phytate content in the final product to a nutritionally acceptable level. Optimal conditions for the endogenous phytases of black beans, this is 60C and pH 6.0, resulted in a 55% reduction in 1P₆ after soaking and cooking. the sum of 1P5 and 1P₅ was reduced by 54%. 1P₆ reduction was most extensive when black beans were soaked at 50C while adding exogenous phytases during the last 2 h of soaking. After soaking and cooking 1P₆ was degraded by 85% and the sum of 1P6 and 1P5 was reduced by 82 % compared to the values in raw beans when adding Escherichia coli phytase. Using rye phytase the values were estimated to be 73% and 70%, thereby a clear accumulation of IP₄ occurred. Thus, a significant improvement in reduction of mnyo-inositol phosphates with adverse effects on mineral bioavailability has been achieved in comparison to the usual household procedure.     

PURIFICATION AND CHARACTERIZATION OF A PHYTASE FROM SPELT

Four soluble phytases were identified in germinating spelt. Although numerous purification strategies were applied, none of the four phytases could be purified to homogeneity. The purest phytase preparation, called D21, contained a phytase (major component) and an acid phosphatase (APH) (minor component). The phytase behaves like a monomeric protein of a molecular mass of about 68 kDa and shows a broad substrate specificity. Optimal pH for degradation of phytate was 6.0 and the optimal temperature 45C. Kinetic parameters for the hydrolysis of Na-phytate were K_M 400 μmoll^?1 and k_cat 368s^?1 at pH 6.0. The spelt phytase D21 degrades phytate stepwise.     

ENDOGENOUS PHYTATE-DEGRADING ENZYMES ARE RESPONSIBLE FOR PHYTATE REDUCTION WHILE PREPARING BEANS (PHASEOLUS VULGARIS)

The three dry nonprocessed bean varieties contain high amounts of phytate ranging from 9.3 to 15.9 μmol g^-1 dry basis, representing 85 to 89% of total myo-inositol phosphates, while the other myo-inositol phosphates were found only in small or trace amounts. Myo-inositol phosphate concentrations were not affected by soaking in water for 15h at 25C, whereas cooking resulted in a significant reduction in phytate content (16 to 24%) with a concomitant increase in the concentrations of the lower myo-inositol phosphates. The sum of phytate and myo-inositol pentakisphosphate after soaking and cooking represents about 93% of the amount in raw beans. Therefore, preparing of beans has only a limited effect on the content of myo-inositol phosphates with inhibitory effects on mineral absorption. Phytate-degrading enzymes (phytases) were identified as responsible for phytate removal during cooking, since a good correlation between phytase activity and phytate hydrolysis was observed.     

The Nucleotide Sequence of a 39 kb Segment of Yeast Chromosome IV: 12 New Open Reading Frames,Nine Known Genes and One Gene for Gly-tRNA

The complete nucleotide sequence of a 39 090 bp segment from the left arm of yeast chromosome IV was determined. Twenty-one open reading frames (ORFs) longer than 100 amino acids and a Gly-tRNA gene were discovered. Nine of the 21 ORFs (D0892, D1022, D1037, D1045, D1057, D1204, D1209, D1214, D1219) correspond to the previously sequenced Saccharomyces cerevisiae genes for the NAD-dependent glutamate dehydrogenase (GDH), the secretory component (SHR3), the GABA transport protein (UGA4), the high mobility group-like protein (NHP2), the hydroxymethylbilane synthase (HEM3), the methylated DNA protein-cysteine S-methyltransferase (MGT1), a putative sugar transport protein, the Shm1 protein (SHM1) and the anti-silencing protein (ASF2). The inferred amino acid sequences of 11 ORFs show significant similarity with known proteins from various organisms, whereas the remaining ORF does not share any similarity with known proteins. The nucleotide sequence has been entered in the EMBL data Library under the Accession Number X99000.©1997 John Wiley & Sons, Ltd.