scholarly journals Cloning, expression and characterization of l-asparaginase from Pseudomonas fluorescens for large scale production in E. coli BL21

3 Biotech ◽  
2015 ◽  
Vol 5 (6) ◽  
pp. 975-981 ◽  
Author(s):  
Vijay Kishore ◽  
K. P. Nishita ◽  
H. K. Manonmani
Keyword(s):  
Pseudomonas Fluorescens ◽  
Large Scale ◽  
Scale Production ◽  
E Coli ◽  
Large Scale Production

scholarly journals Cloning, expression, and analysis of the group 2 allergen from Dermatophagoides farinae from China

2010 ◽  
Vol 82 (4) ◽  
pp. 941-951 ◽  
Author(s):  
Cui Yu-bao ◽  
Ying Zhou ◽  
Shi Weihong ◽  
Ma Guifang ◽  
Li Yang ◽  
...  
Keyword(s):  
Large Scale ◽  
Alpha Helix ◽  
Random Coil ◽  
Reference Sequence ◽  
Scale Production ◽  
Dermatophagoides Farinae ◽  
E Coli ◽  
Large Scale Production ◽  
Solid Foundation ◽  
Group 2

To obtain the recombinant group 2 allergen product of Dermatophagoides farinae (Der f 2), the Der f 2 gene was synthesized by RT-PCR. The full-length cDNA comprised 441 nucleotides and was 99.3% identical to the reference sequence (GenBank AB195580). The cDNA was bound to vector pET28a to construct plasmid pET28a(+)-Der f 2, which was transformed into E. coli BL21 and induced by IPTG. SDS-PAGE showed a specific band of about 14kDa in the hole cell lysate. s estiated by chroatography, about 3.86 g of the recobinant product as obtained, which conjugated with serum IgE from asthmatic children. The protein had a signal peptide of 17 amino acids. Its secondary structure comprised an alpha helix (19.86%), an extended strand (30.82%), and a random coil (49.32%). The subcellular localization of this allergen was predicted to be at mitochondria. Furthermore, its function was shown to be associated with an MD-2-related lipid-recognition (ML) domain. The results of this study provide a solid foundation for large-scale production of the allergen for clinical diagnosis and treatent of allergic disorders.

scholarly journals Large-scale production and physicochemical characterization of human immune interferon.

1979 ◽  
Vol 26 (1) ◽  
pp. 36-41 ◽  
Author(s):  
M P Langford ◽  
J A Georgiades ◽  
G J Stanton ◽  
F Dianzani ◽  
H M Johnson
Keyword(s):  
Large Scale ◽  
Physicochemical Characterization ◽  
Scale Production ◽  
Immune Interferon ◽  
Large Scale Production ◽  
Human Immune

scholarly journals Advances in the Metabolic Engineering of Escherichia coli for the Manufacture of Monoterpenes

Catalysts ◽  
2019 ◽  
Vol 9 (5) ◽  
pp. 433 ◽  
Author(s):  
Si-si Xie ◽  
Lingyun Zhu ◽  
Xin-yuan Qiu ◽  
Chu-shu Zhu ◽  
Lv-yun Zhu
Keyword(s):  
Large Scale ◽  
Metabolic Flux ◽  
Process Development ◽  
Pathway Engineering ◽  
Scale Production ◽  
Future Studies ◽  
E Coli ◽  
Large Scale Production ◽  
Rate Limiting ◽  
Dependent Pathway

Monoterpenes are commonly applied as pharmaceuticals and valuable chemicals in various areas. The bioproduction of valuable monoterpenes in prokaryotic microbial hosts, such as E. coli, has progressed considerably thanks to the development of different outstanding approaches. However, the large-scale production of monoterpenes still presents considerable limitations. Thus, process development warrants further investigations. This review discusses the endogenous methylerythritol-4-phosphate-dependent pathway engineering and the exogenous mevalonate-dependent isoprenoid pathway introduction, as well as the accompanied optimization of rate-limiting enzymes, metabolic flux, and product toxicity tolerance. We suggest further studies to focus on the development of systematical, integrational, and synthetic biological strategies in light of the inter disciplines at the cutting edge. Our review provides insights into the current advances of monoterpene bioengineering and serves as a reference for future studies to promote the industrial production of valuable monoterpenes.

scholarly journals Preclinical Characterization of Naturally Occurring Polyketide Cyclophilin Inhibitors from the Sanglifehrin Family

2011 ◽  
Vol 55 (5) ◽  
pp. 1975-1981 ◽  
Author(s):  
Matthew A. Gregory ◽  
Michael Bobardt ◽  
Susan Obeid ◽  
Udayan Chatterji ◽  
Nigel J. Coates ◽  
...  
Keyword(s):  
Large Scale ◽  
Scale Production ◽  
Subgenomic Replicon ◽  
Isomerase Activity ◽  
Lead Compounds ◽  
Naturally Occurring ◽  
Large Scale Production ◽  
Pharmacokinetic Properties ◽  
Biosynthetic Engineering

ABSTRACTCyclophilin inhibitors currently in clinical trials for hepatitis C virus (HCV) are all analogues of cyclosporine (CsA). Sanglifehrins are a group of naturally occurring cyclophilin binding polyketides that are structurally distinct from the cyclosporines and are produced by a microorganism amenable to biosynthetic engineering for lead optimization and large-scale production by fermentation. Preclinical characterization of the potential utility of this class of compounds for the treatment of HCV revealed that the natural sanglifehrins A to D are all more potent than CsA at disrupting formation of the NS5A-CypA, -CypB, and -CypD complexes and at inhibition of CypA, CypB, and CypD isomerase activity. In particular, sanglifehrin B (SfB) was 30- to 50-fold more potent at inhibiting the isomerase activity of all Cyps tested than CsA and was also shown to be a more potent inhibitor of the 1b subgenomic replicon (50% effective concentrations [EC50s] of 0.070 μM and 0.16 μM in Huh 5-2 and Huh 9-13 cells, respectively). Physicochemical and mouse pharmacokinetic analyses revealed low oral bioavailability (F< 4%) and low solubility (<25 μM), although the half-lives (t1/2) of SfA and SfB in mouse blood after intravenous (i.v.) dosing were long (t1/2> 5 h). These data demonstrate that naturally occurring sanglifehrins are suitable lead compounds for the development of novel analogues that are less immunosuppressive and that have improved metabolism and pharmacokinetic properties.

cDNA cloning and heterologous expression of a wheat proteinase inhibitor of subtilisin and chymotrypsin (WSCI) that interferes with digestive enzymes of insect pests

2005 ◽  
Vol 386 (4) ◽  
pp. 383-389 ◽  
Author(s):  
Simone Di Gennaro ◽  
Anna G. Ficca ◽  
Daniela Panichi ◽  
Elia Poerio
Keyword(s):  
Proteinase Inhibitor ◽  
Large Scale ◽  
Insect Pests ◽  
Expression System ◽  
Physiological Role ◽  
Scale Production ◽  
Insect Larvae ◽  
E Coli ◽  
Large Scale Production ◽  
Degenerate Oligonucleotide

Abstract A cDNA encoding the proteinase inhibitor WSCI (wheat subtilisin/chymotrypsin inhibitor) was isolated by RT-PCR. Degenerate oligonucleotide primers were designed based on the amino acid sequence of WSCI and on the nucleotide sequence of the two homologous inhibitors (CI-2A and CI-2B) isolated from barley. For large-scale production, wsci cDNA was cloned into the E. coli vector pGEX-2T. The fusion protein GST-WSCI was efficiently produced in the bacterial expression system and, as the native inhibitor, was capable of inhibiting bacterial subtilisin, mammalian chymotrypsins and chymotrypsin-like activities present in crude extracts of a number of insect larvae (Helicoverpa armigera, Plodia interpunctella and Tenebrio molitor). The recombinant protein produced was also able to interfere with chymotrypsin-like activity isolated from immature wheat caryopses. These findings support a physiological role for this inhibitor during grain maturation.

scholarly journals Secretory Expression of YY(3-36) Peptide in E. coli

2019 ◽  
Author(s):  
Sorush Niknamian
Keyword(s):  
Large Scale ◽  
Peptide Yy ◽  
Signal Sequence ◽  
Expression System ◽  
Complete Sequence ◽  
Scale Production ◽  
E Coli ◽  
Large Scale Production ◽  
Human Experiments ◽  
Clinical Experiments

Obesity is the prime suspect in a wide frequency of diabetes type II and cardiovascular diseases worldwide. Recombinant YY (tyrosine-tyrosine) peptide is a locally acting hormone, controlling secretion in the digestive tract. Interestingly, it was later shown that a truncated version of YY peptide, YY(3-36) peptide, has the potential as an important biopharmaceutical in a fight against obesity. This peptide has shown promising results in human clinical experiments in appetite reduction in human experiments. To develop an economical expression system for large-scale production of the peptide in gram-negative bacteria, we have developed a chimeric gene for extracellular expression of this peptide with the assistance of signal sequence of asparaginase II from Escherichia coli. This system has the advantage of producing the complete sequence of YY(3-36) without any extra tags that require further removal with the assistance of expensive specific proteases and reduce the downstream steps significantly. Our results pave the way for the recombinant production of YY(3-36) peptide and further proves the efficacy of asparaginase II signal sequence as a communicator of foreign peptides and proteins into extracellular space of E. coli.

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