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He Lisi, Su Jiale, Liu Xiaoqing, Li Chang and Chen Shangping

irreversible hydrolysis of sucrose into glucose and fructose, are a group of ubiquitous enzymes that can be divided into acid, neutral, or alkaline groups according to their solubility, pH optima, and subcellular localizations. Acid invertase exists in the

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Jun Matsumoto, Hideyuki Goto, Yasutaka Kano, Akira Kikuchi, Hideaki Ueda and Yuta Nakatsubo

growth, it is stored in larger, mature cells. Elevated sucrose level in melon fruit is associated with a decline in acid invertase activity ( Chrost and Schmitz, 1997 ) and an increase in SPS activity ( Gao et al., 1999 ; Hubbard et al., 1989 ; Hubbard

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Anil P. Ranwala and William B. Miller

In mature Lilium longiflorum flower buds, anther and stigma had the highest soluble acid invertase activity [3.29 and 2.31 μmol of reducing sugars (RS)/min per gram of fresh weight (FW), respectively] compared to style, ovary, petal, and filament with activities of 1.52, 1.08, 0.99 and 0.98 μmol RS/min per gram of FW, respectively. DEAE-sephacel chromatography revealed that invertase activity in petal, ovary, style, and stigma was composed exclusively of invertase II and III isoforms. Anther invertase was mainly invertase I with small amounts of invertase II and III. Filament tissue mainly had invertase II and III isoforms with a small amount of invertase I. Wall-bound invertases were extracted with 1.0 m NaCl. Anthers had the highest wall-bound invertase activity (4.42 μmol RS/min per gram of FW) followed by stigma (0.42 μmol RS/min per gram of FW). Other tissues had low wall-bound invertase activity (<0.1 μmol RS/min per gram FW). For further purification, the binding of soluble invertases to nine different reactive dyes was investigated. Invertase I was bound to Reactive Green 5, Reactive Green 19, and Reactive Red 120 columns and was eluted with 0.5 m NaCl, resulting in increase in specific activity ≈10-fold with ≈70% recovery. Invertase II and III bound only to Reactive Red 120. Elution with 0.5 m NaCl resulted in an ≈6-fold increase in specific activity.

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Takaya Moriguchi, Kazuyuki Abe, Tetsuro Sanada and Shohei Yamaki

Soluble sugar content and activities of the sucrose-metabolizing enzymes sucrose synthase (SS) (EC 2.4.1.13), sucrose-phosphate synthase (SPS) (EC 2.4.1.14), and acid invertase (EC 2.4.1.26) were analyzed in the pericarp of fruit from pear cultivars that differed in their potential to accumulate sucrose to identify key enzymes involved in sucrose accumulation in Asian pears. The Japanese pear `Chojuro' [Pyrus pyrifolia (Burro. f.) Nakai] was characterized as a high-sucrose-accumulating type based on the analysis of mature fruit, while the Chinese pear `Yali' (P. bretschneideri Rehd.) was a low-sucrose-accumulating type throughout all developmental stages. The activity of SS and SPS in `Chojuro' increased during maturation concomitant with sucrose accumulation, whereas the activity of these enzymes in `Yali' did not increase during maturation. The activity of SS and SPS in the former were seven and four times, respectively, higher than those in the latter at the mature stage. Further, among 23 pear cultivars, SS activity was closely correlated with sucrose content, while SPS activity was weakly correlated. Soluble acid invertase activity in `Chojuro' and `Yali' decreased with fruit maturation, but the relationships between soluble invertase activity and sucrose content were not significant. The results indicate that SS and SPS are important determinants of sucrose accumulation in Asian pear fruit and that a decrease of soluble acid invertase activity is not absolutely required for sucrose accumulation.

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Gene Lester, Luis Saucedo Arias and Miguel Gomez-Lim

Muskmelon [Cucumis melo L. (Reticulatus Group)] fruit sugar content is the single most important consumer preference attribute. During fruit ripening, sucrose accumulates when soluble acid invertase (AI) activity is less then sucrose phosphate synthase (SPS) activity. To genetically heighten fruit sugar content, knowledge of sugar accumulation during fruit development in conjunction with AI and SPS enzyme activities and their peptide immunodetection profiles is needed. Two netted muskmelon cultivars [`Valley Gold' (VG), a high sugar accumulator, and `North Star' (NS), a low sugar accumulator] with similar maturity indices were assayed for fruit sugars, AI, and SPS activity and immunodetection of AI and SPS polypeptides following 2, 5, 10, 15, 20, 25, 30, 35, and 40 (abscission) days after anthesis (DAA). Both cultivars, grown in spring and fall, showed similar total sugar accumulation profiles. Total sugars increased 1.5 fold, from 2 through 5 DAA and then remained unchanged until 30 DAA. From 30 DAA until abscission, total sugar content increased, with VG accumulating significantly more sugar then NS. In both cultivars, during both seasons, sucrose was detected at 2 DAA, which coincided with higher SPS activity than AI activity. At 5 through 25 DAA, SPS activity was less then AI activity resulting in little or no sucrose detection. It was not until 30 DAA that SPS activity was greater than AI activity resulting in increased sucrose accumulation. VG at abscission had higher total sugar content and SPS activity and lower AI activity than NS. Total polypeptides from both cultivars 2 through 40 DAA, were immunodetected with antibodies: anti-AI and anti-SPS. NS had Al isoforms bands at 75, 52, 38, and 25 kDa that generally decreased wtih DAA. One isoform at 52 kDa remained detectable up to anthesis (40 DAA) VG had the same four Al isoforms, all decreased with DAA and became undetectable by 20 DAA. It is unclear if one or all AI isoforms correspond with detected enzyme activity. VG and NS had one SPS band at 58 kDa that increased with DAA and concomitantly with SPS activity. VG had a more intense SPS polypeptide band at abscission then did NS. Thus, netted muskmelon sugar accumulation may be increased by selecting for cultivars with a specific number of AI isoforms, which are down-regulated, and with high SPS activity during fruit ripening.

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Gene E. Lester, Luis Saucedo Arias and Miguel Gomez-Lim

Muskmelon [Cucumis melo L. (Reticulatus Group)] fruit sugar content is the single most important consumer preference attribute. During fruit ripening, sucrose accumulates when soluble acid invertase (AI) activity is less then sucrose phosphate synthase (SPS) activity. To genetically heighten fruit sugar content, knowledge of sugar accumulation during fruit development in conjunction with AI and SPS enzyme activities and their peptide immunodetection profiles, is needed. Two netted muskmelon cultivars, Valley Gold a high sugar accumulator, and North Star a low sugar accumulator, with identical maturity indices were assayed for fruit sugars, AI and SPS activity, and immunodetection of AI and SPS polypeptides 2, 5, 10, 15, 20, 25, 30, 35, or 40 (abscission) days after anthesis (DAA). Both cultivars, grown in the Fall, 1998 and Spring, 1999, showed similar total sugar accumulation profiles. Total sugars increased 1.5 fold, from 2 through 5 DAA, then remained unchanged until 30 DAA. From 30 DAA until abscission, total sugar content increased, with `Valley Gold' accumulating significantly more than `North Star'. During both seasons, sucrose was detected at 2 DAA, which coincided with SPS activity higher than AI activity, at 5 through 25 DAA, no sucrose was detected which coincided with SPS activity less than AI activity. At 30 DAA when SPS activity was greater than AI activity, increased sucrose accumulation occurred. `Valley Gold' at abscission had higher total sugar content and SPS activity, and lower AI activity than `North Star'. `North Star' had AI isoforms at 75, 52, 38, and 25 kDa (ku) that generally decreased with maturation, although the isoform at 52 ku remained detectable up to anthesis (40 DAA). `Valley Gold' had the same four AI isoforms, all decreased with maturation and became undetectable by 20 DAA. Both `Valley Gold' and `North Star' had one SPS band at 58 ku that increased with DAA, and coincided with SPS activity. `Valley Gold' had a more intense SPS polypeptide band at abscission than `North Star'. Thus, netted muskmelon fruit sugar accumulation may be increased, either by genetic manipulation or by selecting for cultivars with a specific number of down-regulated AI isoforms, and higher SPS activity during fruit ripening.

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Yosef Burger and Arthur A. Schaffer

transition from the stage of fruit growth to that of sucrose accumulation, characterized by a developmental loss of soluble acid invertase (AI) activity ( Hubbard et al., 1989 ; Iwatsubo et al., 1992 ; Lester et al., 2001 ; McCollum, et al., 1988

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Jinmin Fu, Bingru Huang and Jack Fry

, understanding the enzyme activity affecting sucrose metabolism is critical. Sucrose synthesis can be regulated by rapid changes in the activity of sucrose phosphate synthase, sucrose synthase, and acid invertase ( Castrillo, 1992 ; Hawker, 1985 ; Huber and

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Natalie L. Hubbard, D. Mason Pharr and Steven C. Huber

Muskmelon (Cucumis melo L.) fruit lack a stored starch reserve and therefore depend on translocated photoassimilate from the leaf canopy for sugar accumulation during ripening. The influence of canopy photosynthesis on sucrose' accumulation within muskmelon fruit mesocarp was examined. Canopy photosynthetic activities were estimated in a sweet and a nonsweet genotype. Photosynthetic rate of the nonsweet genotype, on a per-plant basis, was only 56% of that of the sweet genotype. The effect of limiting leaf area of the sweet genotype on carbohydrate concentrations and sucrose metabolizing enzymes within the fruit was evaluated. A 50% reduction of leaf area 8 days before initiation of fruit sucrose accumulation resulted in canopy photosynthesis similar to that of the nonsweet genotype. Reduced photosynthetic activity resulted in slightly lower soluble-carbohydrate concentration in the fruit; however, fruit sucrose concentration was three times higher than that reported previously for the nonsweet genotype. The extent to which `fruit sucrose phosphate synthase (SPS) activity increased during maturation was diminished by leaf removal. Acid invertase activity declined in all fruit in a similar manner irrespective of defoliation. A reduction of leaf area of a sweet genotype reduced sucrose accumulation within the fruit. Lower fruit sucrose concentration was associated with lower concentration of raffinose saccharides and lower SPS activity within the fruit. Additionally, insufficient assimilate supply was judged not to be the factor responsible for low sucrose accumulation in a nonsweet genotype.

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Riccardo Lo Bianco and Mark Rieger

The peach [Prunus persica (L.) Batsch (Peach Group)] fruit is a sink organ comprised of different types of tissue, which undergoes three distinct developmental stages during the growth season. The objective of this study was to characterize the activity and partitioning of sorbitol and sucrose catabolism within `Encore' peach fruit to determine whether the two forms of translocated carbon play different roles in the various fruit tissues and/or stages of development. Sorbitol catabolic activity was defined as the sum of NAD-dependent sorbitol dehydrogenase (SDH) and sorbitol oxidase (SOX) activities, whereas sucrose catabolic activity was defined as the sum of sucrose synthase (SS), soluble acid invertase (AI), and neutral invertase (NI) activities. Partitioning of sorbitol and sucrose catabolism in each tissue was calculated as percentage of total sorbitol or sucrose catabolic activity in the entire fruit. At cell division, sorbitol catabolic activity was similar in the endocarp and mesocarp, but lower in the seed. However, sorbitol catabolism was mostly partitioned into the mesocarp, due to its large size compared to that of other tissues. SDH was more active in the mesocarp, while SOX was more active in the endocarp. Sucrose catabolism was most active and partitioned mainly into the endocarp. At endocarp hardening, both sorbitol and sucrose catabolic activities were highest in the seed, but despite this, sucrose catabolism was partitioned mostly in the mesocarp. At cell expansion, sorbitol and sucrose catabolic activities were still higher in the seed only when expressed on a weight basis and similar in mesocarp and seed when expressed on a protein basis. Both sorbitol and sucrose catabolism were partitioned mostly into the mesocarp. Sorbitol and sucrose contents were generally higher in the tissues that exhibited lower catabolic activities. All carbohydrates were always partitioned mostly into the mesocarp. Our results show that, at the cell division and endocarp hardening stages, sorbitol and sucrose catabolism are partitioned differently in the fruit and that SDH activity may play an important role in mesocarp cell division and final fruit size determination.