Showing papers in "Protein Expression and Purification in 1992"

TL;DR: This review covers the principles and practice of IMAC that can be performed under very mild, nondenaturing conditions, and the immobilized metal ion ligand complexes are more likely to withstand wear and tear than are antibodies or enzymes.

TL;DR: The design and use of a bacterial expression vector that contains the signal peptide from the pelB gene of Erwinia carotovora linked to a small peptide epitope for an unusual calcium-dependent antibody (HPC4) permits large-scale expression and purification of recombinant soluble tissue factor and should be generally applicable to the isolation of other recombinant proteins.

TL;DR: A cDNA clone that encodes oryzacystatin, a cysteine protease inhibitor from rice, was isolated and expressed in Escherichia coli BL-21 (DE3) using an expression plasmid under the control of a T7 RNA polymerase promoter.

TL;DR: The modified immunoaffinity column isolation protocol suitable for proteins such as rhMIS is described and the biochemical and antiproliferative properties of this protein are described.

TL;DR: A novel plasmid expression vector (pH6EX3) that directs the synthesis of a fusion protein with a histidine hexapeptide at its N-terminus and a foreign protein at its C-terminal was constructed.

TL;DR: A cDNA clone, lambda GTHA1, encoding human glutathione transferase A1-1 has been isolated from a hepatoma HepG2 cDNA library, showing minor differences from cDNA deriving from normal liver, but the deduced amino acid sequence was identical to the structure previously described.

TL;DR: P-Hydroxyacetophenone was coupled to epoxy-activated Sepharose 6B to generate an affinity chromatographic matrix to purify aldehyde dehydrogenase and was found to be a competitive inhibitor against propionaldehyde and noncompetitive against NAD.

TL;DR: The recombinant tyrosine-ester sulfotransferase was identified by its unique substrate spectrum, by comparison with three peptides that were sequenced from homogeneous tyrosines isolated directly from rat liver, and by the specificity of antibody raised to the rat liver enzyme.

TL;DR: The purification procedures reported here for recombinant and native calreticulin yield homogeneous preparations of the proteins, as judged by the HPLC reverse-phase chromatography and by sodium dodecyl sulfate-polyacrylamide gel electrophoresis.

TL;DR: N-Ser PLA2 demonstrated the same kinetics of activation as the wild type enzyme on large vesicles of zwitterionic lipid and showed the same specific activity as thewild-type protein.

TL;DR: An improved method for isolating SGT from greening potato peel using two new chromatographic supports, Macro-Prep 50 Q anion-exchange and Superdex 75HR size exclusion media, under medium-pressure conditions at room temperature is described.

TL;DR: Two recombinant baculoviruses that express the alpha and beta subunits of Drosophila melanogaster casein kinase II, respectively, have been constructed.

TL;DR: The gene for the indole alkaloid biosynthetic enzyme, strictosidine synthase, of Catharanthus roseus has been cloned into an inducible Escherichia coli expression vector using an expression cassette polymerase chain reaction technique.

TL;DR: General procedures for the rapid, large-scale purification of recombinant Lactobacillus casei thymidylate synthase and its mutants have been established and an effective method employs sequential phosphocellulose and hydroxylapatite chromatography.

TL;DR: An inducible, high-copy plasmid for the expression of foreign proteins in Escherichia coli, pPB1, which combines the trc promoter, beta-galactosidase translation start site, and polylinker of pKK233-2 with the origin of replication region of pUC19 is reported.

TL;DR: Investigation of immunoaffinity-purified recombinant human asparagine synthetase in yeast minimal medium demonstrated both glutamine-dependent and ammonia-dependentAsparagine Synthetase activities, as well as glutaminase activity, which was significantly higher than that of a previously reported yeast expression system.

TL;DR: It is concluded from these studies that tailoring recombinant proteins with three or more alternate histidines should result in the isolation of such chimeric proteins from crude mixtures in a single step.

TL;DR: The purified ET receptor was active in ET binding and was resolved by sodium dodecyl sulfate-polyacrylamide gel electrophoresis into two polypeptides with apparent molecular masses of 45 and 35 kDa, indicating that the NH2-terminal 64-amino-acid residues are not essential for ligand binding.

TL;DR: The results suggest that purification by crystallization may be useful for other overexpressed enzymes and multienzymes complexes and for investigations of other properties of the separate subunits.

TL;DR: It was found that, contrary to the way yeast modifies its own cytochromes c, the recombinant proteins were partially acetylated at their N-terminus, except for the drosophila protein, which remained entirely unblocked.

TL;DR: This method allows the simple purification of directly expressed Fab' or F(ab')2 fragments from CHO cell supernatant using protein G-Sepharose.

TL;DR: It is concluded that this recombinant rat phenylalanine hydroxylase is essentially identical to the native enzyme and should be very useful in the future study of this important hydroxyase.

TL;DR: In this system, hPDGF-B is expressed as an insoluble, intracellular protein and can readily be obtained in a partially purified form after differential centrifugation and amino acid sequence determination of the purified protein has verified that the amino-terminal portion of the recombinant PDGF is correct.

TL;DR: The glutamyl-tRNA synthetase of Escherichia coli was purified to homogeneity from the overproducing strain DH5 alpha(pLQ7612) by a two-step procedure that takes only about 6 h and yields 10 mg of enzyme per gram of wet cells.

TL;DR: It is demonstrated that Baculovirus-infected Sf9 cells can be used for high-level expression of pancreatic cholesterol esterase and the recombinant enzyme will be useful for further characterization of this protein.

TL;DR: Secretion from recombinant yeast was used as a potential source of large quantities of the leech protein antistasin (ATS), a potent and highly specific inhibitor of the serine protease coagulation factor Xa, and preliminary in vitro characterization indicated that the yeast r-ATS forms are indistinguishable from each other as well as from R-ATS expressed by the insect baculovirus host-vector system.

TL;DR: The purification has been successfully applied to pyruvate decarboxylase isolated from two different species of yeast and therefore is likely to be of general utility for purification of this enzyme from other yeast sources.

TL;DR: Although devoid of measurable proteolytic activity, factor rXai competitively inhibited plasma factor Xa assembly into functional prothrombinase complexes and inhibited plasma clotting in a dose-dependent manner.

TL;DR: The Escherichia coli gene which encodes N-acetylneuraminic acid aldolase was isolated by the polymerase chain reaction, cloned into the inducible expression vector pTTQ18, and overexpressed in E. coli to achieve a high yield and high activity.

TL;DR: Results indicate that the baculovirus/insect cell expression system is well suited for producing alpha 315, a structurally simplified model of the type II calmodulin-dependent protein kinase.