scholarly journals A Seven-Gene Locus for Synthesis of Phenazine-1-Carboxylic Acid by Pseudomonas fluorescens2-79

1998 ◽  
Vol 180 (9) ◽  
pp. 2541-2548 ◽  
Author(s):  
Dmitri V. Mavrodi ◽  
Vladimir N. Ksenzenko ◽  
Robert F. Bonsall ◽  
R. James Cook ◽  
Alexander M. Boronin ◽  
...  
Keyword(s):  
Carboxylic Acid ◽  
Gene Locus ◽  
Acid Metabolism ◽  
Shikimic Acid ◽  
Expression System ◽  
Multienzyme Complex ◽  
Broad Spectrum Antibiotic ◽  
T7 Promoter ◽  
New Genes ◽  
Chorismic Acid

ABSTRACT Pseudomonas fluorescens 2-79 produces the broad-spectrum antibiotic phenazine-1-carboxylic acid (PCA), which is active against a variety of fungal root pathogens. In this study, seven genes designated phzABCDEFG that are sufficient for synthesis of PCA were localized within a 6.8-kbBglII-XbaI fragment from the phenazine biosynthesis locus of strain 2-79. Polypeptides corresponding to allphz genes were identified by analysis of recombinant plasmids in a T7 promoter/polymerase expression system. Products of thephzC, phzD, and phzE genes have similarities to enzymes of shikimic acid and chorismic acid metabolism and, together with PhzF, are absolutely necessary for PCA production. PhzG is similar to pyridoxamine-5′-phosphate oxidases and probably is a source of cofactor for the PCA-synthesizing enzyme(s). Products of thephzA and phzB genes are highly homologous to each other and may be involved in stabilization of a putative PCA-synthesizing multienzyme complex. Two new genes, phzXand phzY, that are homologous to phzA andphzB, respectively, were cloned and sequenced from P. aureofaciens 30-84, which produces PCA, 2-hydroxyphenazine-1-carboxylic acid, and 2-hydroxyphenazine. Based on functional analysis of the phz genes from strains 2-79 and 30-84, we postulate that different species of fluorescent pseudomonads have similar genetic systems that confer the ability to synthesize PCA.

Investigations of the Shikimic Acid Metabolism in Sphagnum magellanicum during Synthesis of Sphagnorubin Induced by Chilling

1981 ◽  
Vol 176 (8) ◽  
pp. 728-736 ◽  
Author(s):  
Hansjörg Rudolph ◽  
Hans-Jürgen Krause ◽  
Michael Blaicher ◽  
Elisabeth Herms
Keyword(s):  
Acid Metabolism ◽  
Shikimic Acid ◽  
Sphagnum Magellanicum

scholarly journals Short-Term Effects of γ-Irradiation on 1-Aminocyclopropane-1-Carboxylic Acid Metabolism in Early Climacteric Cherry Tomatoes

1990 ◽  
Vol 92 (3) ◽  
pp. 577-581 ◽  
Author(s):  
Christian Larrigaudière ◽  
Alain Latché ◽  
Jean Claude Pech ◽  
Christian Triantaphylidès
Keyword(s):  
Carboxylic Acid ◽  
Acid Metabolism ◽  
Short Term ◽  
Γ Irradiation ◽  
Cherry Tomatoes ◽  
Short Term Effects

Analysis of p53 expression in human tumours: an antibody raised against human p53 expressed in Escherichia coli

1992 ◽  
Vol 101 (1) ◽  
pp. 183-189 ◽  
Author(s):  
C.A. Midgley ◽  
C.J. Fisher ◽  
J. Bartek ◽  
B. Vojtesek ◽  
D. Lane ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Polyclonal Antibodies ◽  
P53 Protein ◽  
Expression System ◽  
High Titre ◽  
T7 Promoter ◽  
Aberrant Expression ◽  
P53 Expression ◽  
Normal Human ◽  
Very High

A cDNA encoding the complete normal human p53 protein was expressed in Escherichia coli using an expression system based on the bacteriophage T7 promoter. The cDNA was adapted so that the full-length protein was produced without fusion to any other sequence. Large amounts of the protein were isolated and the purified protein used to produce very high titre polyclonal antibodies to p53. These new antibodies permit the sensitive detection of p53 and p53 complexes in ELISA and immunoblotting assays. Most importantly, they also permit the detection of p53 in archival tumour material that has been conventionally fixed in formalin and embedded in paraffin wax. Using this reagent we have found that aberrant expression of p53 is a frequent feature of human breast cancer. We are able to recognise six different classes of p53 expression pattern that may be of help in the subclassification of breast tumours.

scholarly journals Development of a Dual-Vector System Utilizing MicroRNA Mimics of the Autographa californica miR-1 for an Inducible Knockdown in Insect Cells

2019 ◽  
Vol 20 (3) ◽  
pp. 533
Author(s):  
Krisztina Koczka ◽  
Wolfgang Ernst ◽  
Dieter Palmberger ◽  
Miriam Klausberger ◽  
Lisa Nika ◽  
...  
Keyword(s):  
Fluorescence Intensity ◽  
Insect Cell ◽  
Fluorescent Protein ◽  
Expression System ◽  
Yellow Fluorescent Protein ◽  
Mrna Level ◽  
Vector System ◽  
Sf9 Cells ◽  
Artificial Microrna ◽  
T7 Promoter

The baculovirus-insect cell expression system is a popular tool for the manufacturing of various attractive recombinant products. Over the years, several attempts have been made to engineer and further improve this production platform by targeting host or baculoviral genes by RNA interference. In this study, an inducible knockdown system was established in insect (Sf9) cells by combining an artificial microRNA precursor mimic of baculoviral origin and the bacteriophage T7 transcription machinery. Four structurally different artificial precursor constructs were created and tested in a screening assay. The most efficient artificial microRNA construct resulted in a 69% reduction in the fluorescence intensity of the target enhanced yellow fluorescent protein (eYFP). Next, recombinant baculoviruses were created carrying either the selected artificial precursor mimic under the transcriptional control of the T7 promoter or solely the T7 RNA polymerase under a baculoviral promoter. Upon co-infecting Sf9 cells with these two viruses, the fluorescence intensity of eYFP was suppressed by ~30–40% on the protein level. The reduction in the target mRNA level was demonstrated with real-time quantitative PCR. The presented inducible knockdown system may serve as an important and valuable tool for basic baculovirus-insect cell research and for the improvement of production processes using this platform.

Harvest maturity related changes in the cold-induced activation of 1-aminocyclopropane-1-carboxylic acid metabolism in Granny Smith apples / Efecto del estado de madurez sobre la activación por frío del metabolismo del ácido 1-aminociclopropano-1-carboxílico en manzanas Granny Smith

1999 ◽  
Vol 5 (3) ◽  
pp. 223-228 ◽  
Author(s):  
C. Larrigaudiere ◽  
I. Recasens ◽  
J. Graell ◽  
M. Vendrell
Keyword(s):  
Carboxylic Acid ◽  
Ethylene Production ◽  
Oxidase Activity ◽  
Acid Metabolism ◽  
Cold Treatment ◽  
Acc Oxidase ◽  
Malus Domestica Borkh ◽  
Harvest Dates ◽  
Harmonization Process ◽  
Harvest Maturity

Changes in 1-aminocyclopropane-1-carboxylic acid metabolism in apples ( Malus domestica Borkh cv Granny Smith) were studied in relation to cold storage. Emphasis was given to the differential re sponsiveness of fruits to cold treatment as a function of stage of maturity at harvest. Fruits were held at 1 or 20 °C for 30 days, respectively, or exposed to 1 °C for 10 days and then storaged at 20 °C for up to 30 days. Fruits at 20 °C showed typical climacteric behavior. Differences at 1 °C between maturity stages in ethylene production and ACC oxidase activity were abolished, which showed that cold treatment is an important inducer of climacteric rise in preclimacteric Granny Smith apples. At 1 °C, ethylene production was lower than at 20 °C and the maxima in production were similar for all the stages of maturity, but took place at different times which corresponded exactly to the initial differ ences in harvest dates. After the transfer to 20 °C, fruits exhibited similar behavior as regards ethyl ene production, ACC oxidase activity, and ACC and MACC levels in relation to a harmonization process which is discussed in this study.

The shikimate pathway. Part 8. Synthesis of (−)-3(R)-amino-4(R),5(R)-dihydroxy-1-cyclohexene-1-carboxylic acid: The 3(R)-amino analogue of (−)-shikimic acid

Tetrahedron ◽  
1996 ◽  
Vol 52 (25) ◽  
pp. 8565-8580 ◽  
Author(s):  
Harry Adams ◽  
Neil A. Bailey ◽  
Roger Brettle ◽  
Richard Cross ◽  
Martyn Frederickson ◽  
...  
Keyword(s):  
Carboxylic Acid ◽  
Shikimic Acid ◽  
Shikimate Pathway

A T7 promoter-specific, inducible protein expression system for

1996 ◽  
Vol 250 (2) ◽  
pp. 230 ◽  
Author(s):  
Birgit Conrad ◽  
R. S. Savchenko ◽  
Roland Breves ◽  
J. Hofemeister
Keyword(s):  
Protein Expression ◽  
Expression System ◽  
T7 Promoter ◽  
Protein Expression System ◽  
Inducible Protein

scholarly journals Overproduction and Biochemical Characterization of the Chryseobacterium meningosepticum BlaB Metallo-β-Lactamase

2002 ◽  
Vol 46 (6) ◽  
pp. 1921-1927 ◽  
Author(s):  
Sandrine Vessillier ◽  
Jean-Denis Docquier ◽  
Sandrine Rival ◽  
Jean-Marie Frere ◽  
Moreno Galleni ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Ion Exchange ◽  
Kinetic Parameters ◽  
Culture Supernatant ◽  
Biochemical Characterization ◽  
Expression System ◽  
T7 Promoter ◽  
Hydrolysis Of

ABSTRACT The BlaB metallo-β-lactamase of Chryseobacterium meningosepticum CCUG4310 was overproduced in Escherichia coli by means of a T7 promoter-based expression system. The overproducing system, scaled up in a 15-liter fermentor, yielded approximately 10 mg of BlaB protein per liter, mostly released in the culture supernatant. The enzyme was purified by two ion-exchange chromatographic steps with an overall yield of 66%. Analysis of the kinetic parameters revealed efficient activities (k cat/Km ratios of >106 M−1 s−1) toward most penam and carbapenem compounds, with the exception of the 6-α-methoxypenam derivative temocillin and of biapenem, which were poorer substrates. Hydrolysis of cephalosporins was overall less efficient, with a remarkable variability that was largely due to variable affinities of the BlaB enzyme for different compounds. BlaB was also able to hydrolyze serine-β-lactamase inhibitors, including β-iodopenicillanate, sulbactam and, although less efficiently, tazobactam.

ChemInform Abstract: Synthesis of (-)-(3R)-Amino-(4R),(5R)-dihydroxy-1-cyclohexene-1- carboxylic Acid: The (3R)-Amino Analogue of (-)-Shikimic Acid.

ChemInform ◽  
2010 ◽  
Vol 27 (24) ◽  
pp. no-no
Author(s):  
R. BRETTLE ◽  
R. CROSS ◽  
M. FREDERICKSON ◽  
E. HASLAM ◽  
G. M. DAVIES
Keyword(s):  
Carboxylic Acid ◽  
Shikimic Acid

scholarly journals Construction of a Low-Temperature Protein Expression System Using a Cold-Adapted Bacterium, Shewanella sp. Strain Ac10, as the Host

2007 ◽  
Vol 73 (15) ◽  
pp. 4849-4856 ◽  
Author(s):  
Ryoma Miyake ◽  
Jun Kawamoto ◽  
Yun-Lin Wei ◽  
Masanari Kitagawa ◽  
Ikunoshin Kato ◽  
...  
Keyword(s):  
Low Temperature ◽  
Protein Expression ◽  
Recombinant Protein Expression ◽  
Expression System ◽  
Maximum Yield ◽  
Broad Host Range ◽  
Psychrophilic Bacterium ◽  
T7 Promoter ◽  
Cold Adapted ◽  
Protein Expression System

ABSTRACT A recombinant protein expression system working at low temperatures is expected to be useful for the production of thermolabile proteins. We constructed a low-temperature expression system using an Antarctic cold-adapted bacterium, Shewanella sp. strain Ac10, as the host. We evaluated the promoters for proteins abundantly produced at 4°C in this bacterium to express foreign proteins. We used 27 promoters and a broad-host-range vector, pJRD215, to produce β-lactamase in Shewanella sp. strain Ac10. The maximum yield was obtained when the promoter for putative alkyl hydroperoxide reductase (AhpC) was used and the recombinant cells were grown to late stationary phase. The yield was 91 mg/liter of culture at 4°C and 139 mg/liter of culture at 18°C. We used this system to produce putative peptidases, PepF, LAP, and PepQ, and a putative glucosidase, BglA, from a psychrophilic bacterium, Desulfotalea psychrophila DSM12343. We obtained 48, 7.1, 28, and 5.4 mg/liter of culture of these proteins, respectively, in a soluble fraction. The amounts of PepF and PepQ produced by this system were greater than those produced by the Escherichia coli T7 promoter system.

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