Characterization of Pantoea dispersa UQ68J: producer of a highly efficient sucrose isomerase for isomaltulose biosynthesis

Abstract

Aims: Isolation, identification and characterization of a highly efficient isomaltulose producer. Methods and Results: After an enrichment procedure for bacteria likely to metabolize isomaltulose in sucroserich environments, 578 isolates were screened for efficient isomaltulose biosynthesis using an aniline/diphenylamine assay and capillary electrophoresis. An isolate designated UQ68J was exceptionally efficient in sucrose isomerase activity. Conversion of sucrose into isomaltulose by UQ68J (enzyme activity of 90-100 U mg )1 DW) was much faster than the current industrial strain Protaminobacter rubrum CBS574.77 (41-66 U mg )1 DW) or a reference strain of Erwinia rhapontici (0AE3-0AE9 U mg )1 DW). Maximum yield of isomaltulose at 78-80% of supplied sucrose was achieved in less than half the reaction time needed by CBS574.77, and the amount of contaminating trehalulose (4%) was the lowest recorded from an isomaltulose-producing microbe. UQ68J is a Gram negative, facultatively anaerobic, motile, noncapsulate, straight rod-shaped bacterium producing acid but no gas from glucose. Based on 16S rDNA analysis UQ68J is closest to Klebsiella oxytoca, but it differs from Klebsiella in defining characteristics and most closely resembles Pantoea dispersa in phenotype.Significance and Impact of Study: This organism is likely to have substantial advantage over previously characterized sucrose isomerase producers for the industrial production of isomaltulose.

High-Throughput Profiling of the Fiber and Sugar Composition of Sugarcane Biomass

“…analytical anion exchange column and calibration with sugar standards as described by Wu and Birch [24]. Sugar standards used for calibration were in a dilution series ranging 1-150, 3-300, and 1-150 mM for sucrose, fructose, and glucose, respectively.…”

“…Additionally, fibrated cane (whole stalk) is analyzed in the field environment with SpectraCane (Biolab, Australia), an integrated at-line analyzer comprising a shredder to homogenize cane stalks into a fiber, conveyer, and sample presentation component, and an NIR spectroscopic instrument, as described in [21,22]. High-performance liquid chromatography (HPLC) is a well-established system for quantifying and profiling sugars in sugarcane juice [23,24]. Combination of the juice characterization, provided by HPLC, and fiber characterization, provided by NIR spectroscopy, gives a comprehensive profile of all phenotypic factors contributing to the total biomass for genetic studies.…”

A New Colorimetric Method for Determining the Isomerization Activity of Sucrose Isomerase

“…The enzymatic properties of several microorganisms containing sucrose isomerases, such as Erwinia rhapontici, 8) Klebsiella sp. LX3, 7) Pantoea dispersa UQ68J, 9) and Serratia plymuthica, 10) have been extensively characterized to date.…”

Analysis of genes controlling biomass traits in the genome of sugarcane (Saccharum spp. hybrids)

“…The final diluted solution (1 mL) was transferred to a 96-well plate and subjected to analysis using a Dionex BioLC system DX 600 (Dionex, Sunnyvale, CA, U.S). Sucrose, glucose and fructose concentrations were measured at high pH (85 mM L −1 KOH), with a PA20 analytical anion exchange column and calibration with sugar standards, as described in Wu and Birch (2004). The sugar standards used for calibration were in a dilution series ranging from 1 -150 mM, 3 -300 mM and 1 -150 mM for sucrose, fructose and glucose, respectively.…”

“…High performance liquid chromatography (HPLC) is a well-established system for quantifying and profiling sugars in sugarcane juice Birch, 2004, Wu andBirch, 2007). The combination of the juice characterization, provided by HPLC, and fiber characterization, provided by NIR spectroscopy, gives a comprehensive profile of all phenotypic factors contributing to the total biomass for genetic studies.…”