Molecular cloning and functional identification of invertase isozymes from green bamboo Bambusa oldhamii.
Three Bo beta fruct 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, Bo beta fruct1 and Bo beta fruct2 were determined to encode cell wall invertases, whereas Bo beta fruct3encodes a vacuolar invertase. The recombinant proteins encoded by Bo beta fruct2 and Bo beta fruct3 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 Bo beta fruct1 and Bo beta fruct2 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 Bo beta fruct2-encoded enzyme is important for maintaining the sucrose gradient between source and sink organs, while the predominant expression of Bo beta fruct3 in regions of active cell differentiation and expansion suggests functions in osmoregulation and cell enlargement.
Hsieh
Chang-Wen
CW
Institute of Microbiology and Biochemistry, National Taiwan University, Taipei 106, Taiwan.
Liu
Li-Ka
LK
Yeh
Sheng-Hsiung
SH
Chen
Chi-Fu
CF
Lin
Hsin-I
HI
Sung
Hsien-Yi
HY
Wang
Ai-Yu
AY
D000595:Amino Acid Sequence; D031723:Bambusa / Q000201:enzymology*; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D015870:Gene Expression; D007527:Isoenzymes / Q000032:analysis* / Q000737:chemistry / Q000235:genetics*; D008969:Molecular Sequence Data; D010802:Phylogeny; D010843:Pichia / Q000235:genetics; D011994:Recombinant Proteins; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016415:Sequence Alignment; D043324:beta-Fructofuranosidase / Q000032:analysis* / Q000737:chemistry / Q000235:genetics*
D018076:DNA, Complementary; D007527:Isoenzymes; D011994:Recombinant Proteins; D043324:beta-Fructofuranosidase