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Score: 1.00 | | Journal: Protein Expr . Purif . | Literature: oryza Field: abstract Doc ID: pub10024476 | Abstract: A putative rice trypsin/chymotrypsin inhibitor of the Bowman-Birk family , RBBI-8 of about 20 kDa , was expressed in Escherichia coli as a fusion protein bearing an N-terminal ( His ) 6 purification tag .
The expressed recombinant protein , rRBBI-8 , is insoluble and accumulates as inclusion bodies .
The insoluble protein was solubilized in 8 M urea under reducing environment and then refolded into its active conformation under optimized redox conditions .
Strategies used to optimize yield and efficiency include selecting the redox system , increasing protein concentration during refolding by adding the denatured protein in a stepwise way , utilizing additives to prevent aggregation , and selecting buffer-exchanging conditions .
A Ni-chelate affinity column was then employed to purify the renatured protein . rRBBI-8 shows strong inhibitory activity against trypsin and it can slightly inhibit chymotrypsin .
In this study , a refolding and purification system was set up for this cysteine-rich recombinant protein expressed in a prokaryotic system . | Supplemental links/files: reference in endnote online text related articles pubmed citation | |
Score: 2.00 | | Journal: Biochemistry | Literature: oryza Field: abstract Doc ID: pub10029544 | Abstract: The Na , K-ATPase is specifically inhibited by the cardiac glycoside , ouabain .
Via a largely undefined mechanism , the ouabain affinity of the Na , K-ATPase can be manipulated by mutating the residues at the borders of the first extracellular ( M1-M2 ) loop of the alpha subunit [ Price , E M , Rice , D A , and Lingrel , J B ( 1990 ) J Biol . Chem . 265 , 6638-6641 ] .
To address this issue , we compared the effects of two combinations of charged residues at the M1-M2 loop border , R113 , D124 and D113 , R124 ( numbered according to the rat alpha1 subunit ) , on the ouabain sensitivity of the alpha1 and alpha2 isoforms .
We report that ouabain sensitivity is dependent not only upon the identity of the residues at the M1-M2 loop border but also upon the context into which they are introduced .
Furthermore , at low concentrations of ATP , the identity of the residues at the M1-M2 loop border affects the regulation of ATP hydrolysis by potassium in an isoform-specific manner .
Analysis of chimeric alpha subunits reveals that the effects of potassium are determined primarily by the interaction of the N-terminus and M1-M2 loop with the C-terminal third of the alpha subunit .
M1-M2 loop border residues may , therefore , influence ouabain sensitivity indirectly by altering the stability or structure of the intermediate of the Na , K-ATPase catalytic cycle which is competent to bind ouabain .
| Supplemental links/files: reference in endnote online text related articles pubmed citation | |
Score: 1.00 | | Journal: Plant Cell Physiol .
| Literature: oryza Field: abstract Doc ID: pub10050310 | Abstract: Recent evidence has shown that the prolamine polysomes are attached not only to the endoplasmic reticulum membranes that bound the prolamine protein bodies ( PBs ) but also to cytoskeleton elements associated with this subcellular fraction .
To learn more about the nature of the proteins that are associated with this supra-macromolecular complex , proteins extracted from an enriched cytoskeleton-PB fraction were resolved by two-dimensional polyacrylamide gel electrophoresis under non-equilibrium conditions and analyzed for their composition by immunological and biochemical methods .
Immunoblot analysis indicated the presence of the cytoskeletal proteins , actin and tubulin , and the cytoskeletal-associated protein EF1 alpha in this fraction .
Microsequencing of selected polypeptides revealed a diversity of protein sequences .
In addition to contaminating storage proteins which are selectively solubilized by the isolation procedure , several ribosomal proteins and histone H3 were also identified .
Some of the remaining polypeptides showed partial homology to protein sequences deposited in the database , several of which are cytoskeleton-associated proteins . | Supplemental links/files: reference in endnote online text related articles pubmed citation | |
Score: 1.00 | | Journal: Curr . Opin . Plant Biol .
| Literature: oryza Field: abstract Doc ID: pub10066574 | Abstract: The large number of ESTs generated for Arabidopsis and rice in recent years now act as an important complement to whole genome sequencing projects .
The Arabidopsis Genome Initiative has begun a coordinated effort to sequence the entire genome and , as a result , increasing numbers of large sequence entries can be found in the public databases .
In addition , the mitochondrial genome of Arabidopsis has been completely sequenced .
Genome sequencing studies and the public sequence databases have begun to influence the direction of diverse areas of research from physiology to evolution .
| Supplemental links/files: reference in endnote online text related articles pubmed citation | |
Score: 4.00 | | Journal: J Biol . Chem . | Literature: oryza Field: abstract Doc ID: pub10066741 | Abstract: We completely sequenced 13 , 936 nucleotides ( nt ) of a double-stranded RNA ( dsRNA ) of wild rice ( W-dsRNA ) .
A single long open reading frame ( 13 , 719 nt ) containing the conserved motifs of RNA-dependent RNA polymerase and RNA helicase was located in the coding strand .
The identity between entire nucleotide sequence of W-dsRNA and that of the dsRNA of temperate japonica rice ( J-dsRNA , 13 , 952 nt ) was 75 . 5% .
A site-specific discontinuity ( nick ) was identified at nt 1 , 197 from the 5 end of the coding strand of W-dsRNA .
This nick is also located at nt 1 , 211 from the 5 end in the coding strand of J-dsRNA .
The dsRNA copy number was increased more than 10-fold in pollen grains of both rice plants .
This remarkable increase may be responsible for the highly efficient transmission of J-dsRNA via pollen that we already reported .
J-dsRNA and W-dsRNA were also efficiently transmitted to interspecific F1 hybrids .
Seed-mediated dsRNA transmission to F2 plants was also highly efficient when the maternal parent was wild rice .
The efficiency of dsRNA transmission to F2 plants was reduced when the maternal parent was temperate japonica rice ; however , the reduced rates in F2 plants were returned to high levels in F3 plants . | Supplemental links/files: reference in endnote online text related articles pubmed citation | |