Ribulose-1, 5-bisphosphate carboxylase/oxygenase ( EC 4.1.1.39 ) ( RuBP carboxylase ) is a bifunctional enzyme which catalyzes both carboxylation and oxygenation of ribulose-1, 5-bisphosphate (RuBP). These reactions a...Ribulose-1, 5-bisphosphate carboxylase/oxygenase ( EC 4.1.1.39 ) ( RuBP carboxylase ) is a bifunctional enzyme which catalyzes both carboxylation and oxygenation of ribulose-1, 5-bisphosphate (RuBP). These reactions are the first step in photosynthetic carbon fixation and photorespiration respectively. Because of its key role in the formation of plant yield, many studies on the structure of active-site and catalytic mechanism of this enzyme have展开更多
A structural gene with 385 bp, which codes for the mature small subunit (rbcS) of ribulose-1,5-bi9phosphate carboxylase/oxygenase from tobacco, has been redesigned according to the codon usage of E.coli and chemically...A structural gene with 385 bp, which codes for the mature small subunit (rbcS) of ribulose-1,5-bi9phosphate carboxylase/oxygenase from tobacco, has been redesigned according to the codon usage of E.coli and chemically synthesized. To facilitate the systematic investigation of structure-functional relationship of the rbcS by site-specific mutagenesis, twenty one unique restriction sites distributed throughout the synthetic gene were designed. Gene synthesis was started from chemically synthesizing sixteen oligonucleotides each with 23-66 nucleotides, and these oligonu-cleotides were annealed to form eight duplexes from which 5'- and 3'- two half molecules were formed by stepwise T4 DNA ligase reaction, and then each half molecule was cloned into plasmid pWR13, after that, two half molecules were further confirmed by DNA sequencing, finally both half molecules excised from the cloning plasmid were recombinated to form plasmid pCOTrbcS containing the whole structural gene for tobacco rbcS.展开更多
Ribulose-l,5-bisphosphate carboxylase small subunit gene (rbcS) is present with multi-gene family in plant genome. In Glycine max, the rbcS polypeptide (EC4.1.1.39) is encoded by a gene family containing 4-8 members. ...Ribulose-l,5-bisphosphate carboxylase small subunit gene (rbcS) is present with multi-gene family in plant genome. In Glycine max, the rbcS polypeptide (EC4.1.1.39) is encoded by a gene family containing 4-8 members. Three full-length rbcS cDNA clones were isolated and characterized from soybean seedlings, and both of their nucleotide and amino acid sequences showed high similarity. Differential accumulation of the rbcS mRNA was observed among roots, hypocotyls, cotyledons, epicotyls and leaves. The rbcS genes were up-regulated by various external factors such as salicylic acid (SA), salt stress and drought stress. The expression level of rbcS genes after being treated by 2.0 mmol/L SA and0.4% NaCl, respectively, is 2.5-3.0-fold as high as that of control sample. Moreover, soybean rbcS mRNA was accu-mulated with diurnal variation but easily influenced by light and low temperature.展开更多
[Objective] The aim was to study the cloning and sequence analysis of rbcS gene of wild barley under salt stress. [Method] The tender leaf blade of wild barley under salt stress was taken as the experimental material....[Objective] The aim was to study the cloning and sequence analysis of rbcS gene of wild barley under salt stress. [Method] The tender leaf blade of wild barley under salt stress was taken as the experimental material. The primers were designed according to the homology of rbcS gene sequences of wheat and barely in Genbank; then PCR amplification,recovery,ligation,transformation and sequencing of rbcS gene were carried out. [Result] Two rbcS genes including rbcS1 and rbcS2 with the length of 1 252 and 908 bp respectively were cloned from the barely genome. rbcS1 and rbcS2 were both composed by two exons and one intron. The exons length of the two genes was the same of 525 bp,encoding 174 amino acids,and the homology between them was 96%; however,the intron length of rbcS1 and rbcS2 was 448 and 107 bp respectively.展开更多
Ribulose-1, 5-bisphosphate carboxynase/oxygenase large subunit gene (rbcL) of Jiaozhou Bay phytoplankton was amplified from spring, summer and autumn surface seawater DNAs and cloned respectively. About 50 clones were...Ribulose-1, 5-bisphosphate carboxynase/oxygenase large subunit gene (rbcL) of Jiaozhou Bay phytoplankton was amplified from spring, summer and autumn surface seawater DNAs and cloned respectively. About 50 clones were randomly selected from each library and sequenced. If identical amino acid sequences are considered as the same operational taxonomy unit (OTU), 61 OTUs are identified according to inferred amino acid sequences, among them, 21 from spring seawater, 15 from summer seawater and 25 from autumn seawater. Shannon index calculated from OTU abundances reflects the genetic diversity of a community. The indexes of spring, summer and autumn surface seawater phytoplankton are 2.69, 2.44 and 2.76 respectively, indicating that phytoplankton genetic diversity of autumn seawater is the richest. Seasonal variation ofphytoplankton community is significant; the community compositions of three seasons are almost completely different except for the two OTUs shared by summer and autumn. Surface seawater phytoplankton communities are possibly metacommunities different spatially and temporally.展开更多
Ulva prolifera is a typical green alga in subtidal areas and can grow tremendously fast. A highly efficient Rubisco enzyme which is encoded by Up Rbc L gene may contribute to the rapid growth. In this study, the full-...Ulva prolifera is a typical green alga in subtidal areas and can grow tremendously fast. A highly efficient Rubisco enzyme which is encoded by Up Rbc L gene may contribute to the rapid growth. In this study, the full-length Up Rbc L open reading frame(ORF) was identified, which encoded a protein of 474 amino acids. Phylogenetic analysis of Up Rbc L sequences revealed that Chlorophyta had a closer genetic relationship with higher plants than with Rhodophyta and Phaeophyta. The two distinct residues(aa11 and aa91) were presumed to be unique for Rubisco catalytic activity. The predicted three-dimensional structure showed that one α/β-barrel existed in the C-terminal region, and the sites for Mg^(2+)coordination and CO_2 fixation were also located in this region. Gene expression profile indicated that Up Rbc L was expressed at a higher level under light exposure than in darkness. When the culture temperature reached 35℃, the expression level of Up Rbc L was 2.5-fold lower than at 15℃, and the carboxylase activity exhibited 13.8-fold decrease. Up Rbc L was heterologously expressed in E. coli and was purified by Ni^(2+) affinity chromatography. The physiological and biochemical characterization of recombinant Rubisco will be explored in the future.展开更多
文摘Ribulose-1, 5-bisphosphate carboxylase/oxygenase ( EC 4.1.1.39 ) ( RuBP carboxylase ) is a bifunctional enzyme which catalyzes both carboxylation and oxygenation of ribulose-1, 5-bisphosphate (RuBP). These reactions are the first step in photosynthetic carbon fixation and photorespiration respectively. Because of its key role in the formation of plant yield, many studies on the structure of active-site and catalytic mechanism of this enzyme have
基金Project supported by grants from the High Technology Development Program of China,From the National Key Laboratory of Biooganic and Natural Product Chemistry of China,and from the Laboratory of Computer Chemistry,Chinese Academy of Sciences.
文摘A structural gene with 385 bp, which codes for the mature small subunit (rbcS) of ribulose-1,5-bi9phosphate carboxylase/oxygenase from tobacco, has been redesigned according to the codon usage of E.coli and chemically synthesized. To facilitate the systematic investigation of structure-functional relationship of the rbcS by site-specific mutagenesis, twenty one unique restriction sites distributed throughout the synthetic gene were designed. Gene synthesis was started from chemically synthesizing sixteen oligonucleotides each with 23-66 nucleotides, and these oligonu-cleotides were annealed to form eight duplexes from which 5'- and 3'- two half molecules were formed by stepwise T4 DNA ligase reaction, and then each half molecule was cloned into plasmid pWR13, after that, two half molecules were further confirmed by DNA sequencing, finally both half molecules excised from the cloning plasmid were recombinated to form plasmid pCOTrbcS containing the whole structural gene for tobacco rbcS.
基金This work was supported by the National Key Basic Research Special Funds of China (Grant No. G1998010209).
文摘Ribulose-l,5-bisphosphate carboxylase small subunit gene (rbcS) is present with multi-gene family in plant genome. In Glycine max, the rbcS polypeptide (EC4.1.1.39) is encoded by a gene family containing 4-8 members. Three full-length rbcS cDNA clones were isolated and characterized from soybean seedlings, and both of their nucleotide and amino acid sequences showed high similarity. Differential accumulation of the rbcS mRNA was observed among roots, hypocotyls, cotyledons, epicotyls and leaves. The rbcS genes were up-regulated by various external factors such as salicylic acid (SA), salt stress and drought stress. The expression level of rbcS genes after being treated by 2.0 mmol/L SA and0.4% NaCl, respectively, is 2.5-3.0-fold as high as that of control sample. Moreover, soybean rbcS mRNA was accu-mulated with diurnal variation but easily influenced by light and low temperature.
基金Supported by National Natural Science Foundation of China(30471229 )National High Technology Research and Development Program "863" Project(2008AA10Z224)Students Innovative Experimental Projects in Jilin University (2009C81147)~~
文摘[Objective] The aim was to study the cloning and sequence analysis of rbcS gene of wild barley under salt stress. [Method] The tender leaf blade of wild barley under salt stress was taken as the experimental material. The primers were designed according to the homology of rbcS gene sequences of wheat and barely in Genbank; then PCR amplification,recovery,ligation,transformation and sequencing of rbcS gene were carried out. [Result] Two rbcS genes including rbcS1 and rbcS2 with the length of 1 252 and 908 bp respectively were cloned from the barely genome. rbcS1 and rbcS2 were both composed by two exons and one intron. The exons length of the two genes was the same of 525 bp,encoding 174 amino acids,and the homology between them was 96%; however,the intron length of rbcS1 and rbcS2 was 448 and 107 bp respectively.
基金supported financially by the National Natural Science Foundation of China under contract No.40176028.
文摘Ribulose-1, 5-bisphosphate carboxynase/oxygenase large subunit gene (rbcL) of Jiaozhou Bay phytoplankton was amplified from spring, summer and autumn surface seawater DNAs and cloned respectively. About 50 clones were randomly selected from each library and sequenced. If identical amino acid sequences are considered as the same operational taxonomy unit (OTU), 61 OTUs are identified according to inferred amino acid sequences, among them, 21 from spring seawater, 15 from summer seawater and 25 from autumn seawater. Shannon index calculated from OTU abundances reflects the genetic diversity of a community. The indexes of spring, summer and autumn surface seawater phytoplankton are 2.69, 2.44 and 2.76 respectively, indicating that phytoplankton genetic diversity of autumn seawater is the richest. Seasonal variation ofphytoplankton community is significant; the community compositions of three seasons are almost completely different except for the two OTUs shared by summer and autumn. Surface seawater phytoplankton communities are possibly metacommunities different spatially and temporally.
基金financially supported by the Scientific and Technical Supporting Programs of China(2008 BAC49B01)State Ocean Administration Project(200805069)
文摘Ulva prolifera is a typical green alga in subtidal areas and can grow tremendously fast. A highly efficient Rubisco enzyme which is encoded by Up Rbc L gene may contribute to the rapid growth. In this study, the full-length Up Rbc L open reading frame(ORF) was identified, which encoded a protein of 474 amino acids. Phylogenetic analysis of Up Rbc L sequences revealed that Chlorophyta had a closer genetic relationship with higher plants than with Rhodophyta and Phaeophyta. The two distinct residues(aa11 and aa91) were presumed to be unique for Rubisco catalytic activity. The predicted three-dimensional structure showed that one α/β-barrel existed in the C-terminal region, and the sites for Mg^(2+)coordination and CO_2 fixation were also located in this region. Gene expression profile indicated that Up Rbc L was expressed at a higher level under light exposure than in darkness. When the culture temperature reached 35℃, the expression level of Up Rbc L was 2.5-fold lower than at 15℃, and the carboxylase activity exhibited 13.8-fold decrease. Up Rbc L was heterologously expressed in E. coli and was purified by Ni^(2+) affinity chromatography. The physiological and biochemical characterization of recombinant Rubisco will be explored in the future.
