Molecular Cloning and Functional Identification of Invertase Isozymes from Green Bamboo Bambusa oldhamii

Three Boβfruct cDNAs encoding acid invertases were cloned from shoots of the green bamboo Bambusa oldhamii. On the basis of the amino acid sequences of their products and phylogenetic analyses, Boβfruct1 and Boβfruct2 were determined to encode cell wall invertases, whereas Boβfruct3 encodes a vacuol...

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Bibliographic Details
Published in:Journal of agricultural and food chemistry 2006-04, Vol.54 (8), p.3101-3107
Main Authors: Hsieh, Chang-Wen, Liu, Li-Ka, Yeh, Sheng-Hsiung, Chen, Chi-Fu, Lin, Hsin-I, Sung, Hsien-Yi, Wang, Ai-Yu
Format: Article
Language:English
Subjects:
acidity
Amino Acid Sequence
amino acid sequences
Bambusa - enzymology
Bambusa oldhamii
beta-fructofuranosidase
beta-Fructofuranosidase - analysis
beta-Fructofuranosidase - chemistry
beta-Fructofuranosidase - genetics
Biological and medical sciences
cell walls
Cloning, Molecular
complementary DNA
cytochemistry
DNA, Complementary - genetics
enzyme activity
Food industries
Fundamental and applied biological sciences. Psychology
Gene Expression
isoelectric point
Isoenzymes - analysis
Isoenzymes - chemistry
Isoenzymes - genetics
isozymes
messenger RNA
Molecular Sequence Data
nucleotide sequences
Phylogeny
Pichia - genetics
Recombinant Proteins
Reverse Transcriptase Polymerase Chain Reaction
Sequence Alignment
shoots
substrate specificity
vacuoles
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Summary:Three Boβfruct cDNAs encoding acid invertases were cloned from shoots of the green bamboo Bambusa oldhamii. On the basis of the amino acid sequences of their products and phylogenetic analyses, Boβfruct1 and Boβfruct2 were determined to encode cell wall invertases, whereas Boβfruct3 encodes a vacuolar invertase. The recombinant proteins encoded by Boβfruct2 and Boβfruct3 were produced in Pichia pastoris and purified to near homogeneity using ammonium sulfate fractionation and immobilized metal affinity chromatography. The pH optima, pI values, and substrate specificities of the isolated enzymes were consistent with those of plant cell wall or vacuolar invertases. The growth-dependent expression of Boβfruct1 and Boβfruct2 in the base regions of shoots underscores their roles in sucrose unloading and providing substrates for shoot growth. Its high sucrose affinity suggests that the Boβfruct2-encoded enzyme is important for maintaining the sucrose gradient between source and sink organs, while the predominant expression of Boβfruct3 in regions of active cell differentiation and expansion suggests functions in osmoregulation and cell enlargement. Keywords: Bamboo (Bambusa oldhamii); cell-wall invertase; vacuolar invertase; cDNA cloning; recombinant invertase; gene expression
ISSN:0021-8561
1520-5118
DOI:10.1021/jf052711s