scholarly journals Characterization of the AlkS/PalkB-expression system as an efficient tool for the production of recombinant proteins inEscherichia coli fed-batch fermentations

2006 ◽  
Vol 96 (2) ◽  
pp. 326-336 ◽  
Author(s):  
Stefan Makart ◽  
Matthias Heinemann ◽  
Sven Panke
Keyword(s):  
Recombinant Proteins ◽  
Expression System ◽  
Inescherichia Coli ◽  
Fed Batch ◽  
Efficient Tool

scholarly journals Aggregation-prone peptides modulate interferon gamma functionality in naturally occurring protein nanoparticles

2019 ◽  
Author(s):  
José Vicente Carratalá ◽  
Olivia Cano-Garrido ◽  
Julieta Sánchez ◽  
Cristina Membrado ◽  
Eudald Pérez ◽  
...  
Keyword(s):  
Biological Activity ◽  
Protein Aggregation ◽  
Interferon Gamma ◽  
Recombinant Proteins ◽  
Expression System ◽  
Protein Aggregates ◽  
Naturally Occurring ◽  
Protein Nanoparticles ◽  
Ifn Γ

AbstractEfficient protocols for the production of recombinant proteins are indispensable for the development of the biopharmaceutical sector. Approximately 400 recombinant protein-based biopharmaceuticals have been approved in recent decades, with steady growth projected in the coming years. During the expression of a heterologous gene, the protein quality control network is overcome by the disruption in protein homeostasis, leading to protein aggregation. This phenomenon has been described in all expression systems analyzed to date, including prokaryotic and eukaryotic host cells. These protein aggregates have long been considered inert protein clumps devoid of biological activity and their study has largely been neglected. However, in recent years, the classic view of protein aggregates has completely changed with the recognition that these aggregates are a valuable source of functional recombinant proteins. In this study, bovine interferon-gamma (rBoIFN-γ) was engineered to enhance the formation of protein aggregates by the addition of aggregation-prone peptides (APPs) in the generally recognized as safe (GRAS) bacterial Lactococcus lactis expression system. The L6K2, HALRU and CYOB peptides were selected to assess their intrinsic aggregation capability to nucleate protein aggregation. These APPs enhanced the tendency of the resulting protein to aggregate at the expense of the total protein yield. However, fine physicochemical characterization of the resulting intracellular protein nanoparticles (NPs), the protein released from these protein NPs, and the protein purified from the soluble cell fraction indicated that the compactability of protein conformations is directly related to the biological activity of variants of IFN-γ, which is used here as a model protein with therapeutic potential.ImportanceThe demand for recombinant proteins in the pharmaceutical industry is steadily increasing. Emerging novel protein formulations, including naturally occurring protein NPs, might be an alternative to soluble variants for fine analysis at the biophysical level. Such analyses are important to address safety about biological molecules.This study analyzes the effect of aggregation-prone peptides (APPs) on the improvement of the production of naturally occurring protein nanoparticles (NPs) of interferon gamma (IFN-γ) in the generally recognized as safe (GRAS) Lactococcus lactis expression system. In addition, the fine physico-chemical characterization of the resulting proteins, either obtained from the soluble or insoluble cell fractions, indicates that the selected engineered proteins embedded in the protein NPs show higher compactability than their soluble protein counterparts. Conformational compactability is directly related to the biological performance of the recombinant IFN-γ.

Characterization of a pH-inducible promoter system for high-level expression of recombinant proteins inEscherichia coli

1995 ◽  
Vol 47 (2) ◽  
pp. 186-192 ◽  
Author(s):  
Chih-Hsiung Chou ◽  
Aristos A. Aristidou ◽  
Shi-Yuan Meng ◽  
George N. Bennett ◽  
Ka-Yiu San
Keyword(s):  
Recombinant Proteins ◽  
Inducible Promoter ◽  
Inescherichia Coli ◽  
High Level Expression ◽  
High Level ◽  
Promoter System

Characterization of the growth behavior ofLeishmania tarentolae – a new expression system for recombinant proteins

2007 ◽  
Vol 47 (5) ◽  
pp. 384-393 ◽  
Author(s):  
Claudia Fritsche ◽  
Mandy Sitz ◽  
Norman Weiland ◽  
Reinhard Breitling ◽  
Hans-Dieter Pohl
Keyword(s):  
Recombinant Proteins ◽  
Expression System ◽  
Growth Behavior

scholarly journals Lactococcus lactis as expression host for the biosynthetic incorporation of tryptophan analogues into recombinant proteins

2007 ◽  
Vol 409 (1) ◽  
pp. 193-198 ◽  
Author(s):  
Mohamed El Khattabi ◽  
Maarten L. van Roosmalen ◽  
Dennis Jager ◽  
Heidi Metselaar ◽  
Hjalmar Permentier ◽  
...  
Keyword(s):  
Lactococcus Lactis ◽  
Recombinant Proteins ◽  
Prokaryotic Expression ◽  
Expression System ◽  
Spectroscopic Techniques ◽  
Efficient Tool ◽  
Positive Expression ◽  
Expression Host ◽  
Regulated Expression ◽  
Tryptophan Analogues

Incorporation of Trp (tryptophan) analogues into a protein may facilitate its structural analysis by spectroscopic techniques. Development of a biological system for the biosynthetic incorpor-ation of such analogues into proteins is of considerable importance. The Gram-negative Escherichia coli is the only prokaryotic expression host regularly used for the incorporation of Trp analogues into recombinant proteins. Here, we present the use of the versatile Gram-positive expression host Lactococcus lactis for the incorporation of Trp analogues. The availability of a tightly regulated expression system for this organism, the potential to secrete modified proteins into the growth medium and the construction of the trp-synthetase deletion strain PA1002 of L. lactis rendered this organism potentially an efficient tool for the incorporation of Trp analogues into recombinant proteins. The Trp analogues 7-azatryptophan, 5-fluorotryptophan and 5-hydroxytryptophan were incorporated with efficiencies of >97, >97 and 89% respectively. Interestingly, 5-methylTrp (5-methyltryptophan) could be incorporated with 92% efficiency. Successful biosynthetical incorporation of 5-methylTrp into recombinant proteins has not been reported previously.

scholarly journals Harnessing a Transient Gene Expression System in Nicotiana benthamiana to Explore Plant Agrochemical Transporters

Plants ◽  
2021 ◽  
Vol 10 (3) ◽  
pp. 524
Author(s):  
Bingqi Wu ◽  
Zhiting Chen ◽  
Xiaohui Xu ◽  
Ronghua Chen ◽  
Siwei Wang ◽  
...  
Keyword(s):  
Gene Expression ◽  
Transient Expression ◽  
Nicotiana Benthamiana ◽  
Heterologous Gene Expression ◽  
Expression System ◽  
Functional Characterization ◽  
Transient Gene Expression ◽  
High Mobility ◽  
Gene Expression System

Functional characterization of plant agrichemical transporters provided an opportunity to discover molecules that have a high mobility in plants and have the potential to increase the amount of pesticides reaching damage sites. Agrobacterium-mediated transient expression in tobacco is simple and fast, and its protein expression efficiency is high; this system is generally used to mediate heterologous gene expression. In this article, transient expression of tobacco nicotine uptake permease (NtNUP1) and rice polyamine uptake transporter 1 (OsPUT1) in Nicotiana benthamiana was performed to investigate whether this system is useful as a platform for studying the interactions between plant transporters and pesticides. The results showed that NtNUP1 increases nicotine uptake in N. benthamiana foliar discs and protoplasts, indicating that this transient gene expression system is feasible for studying gene function. Moreover, yeast expression of OsPUT1 apparently increases methomyl uptake. Overall, this method of constructing a transient gene expression system is useful for improving the efficiency of analyzing the functions of plant heterologous transporter-encoding genes and revealed that this system can be further used to study the functions of transporters and pesticides, especially their interactions.

scholarly journals Functional expression of the eukaryotic proton pump rhodopsin OmR2 in Escherichia coli and its photochemical characterization

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Masuzu Kikuchi ◽  
Keiichi Kojima ◽  
Shin Nakao ◽  
Susumu Yoshizawa ◽  
Shiho Kawanishi ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Proton Pump ◽  
Expression System ◽  
Spectroscopic Characterization ◽  
Functional Expression ◽  
Proton Pumping ◽  
E Coli ◽  
Oxyrrhis Marina ◽  
Domains Of Life

AbstractMicrobial rhodopsins are photoswitchable seven-transmembrane proteins that are widely distributed in three domains of life, archaea, bacteria and eukarya. Rhodopsins allow the transport of protons outwardly across the membrane and are indispensable for light-energy conversion in microorganisms. Archaeal and bacterial proton pump rhodopsins have been characterized using an Escherichia coli expression system because that enables the rapid production of large amounts of recombinant proteins, whereas no success has been reported for eukaryotic rhodopsins. Here, we report a phylogenetically distinct eukaryotic rhodopsin from the dinoflagellate Oxyrrhis marina (O. marina rhodopsin-2, OmR2) that can be expressed in E. coli cells. E. coli cells harboring the OmR2 gene showed an outward proton-pumping activity, indicating its functional expression. Spectroscopic characterization of the purified OmR2 protein revealed several features as follows: (1) an absorption maximum at 533 nm with all-trans retinal chromophore, (2) the possession of the deprotonated counterion (pKa = 3.0) of the protonated Schiff base and (3) a rapid photocycle through several distinct photointermediates. Those features are similar to those of known eukaryotic proton pump rhodopsins. Our successful characterization of OmR2 expressed in E. coli cells could build a basis for understanding and utilizing eukaryotic rhodopsins.

Spectroscopic Characterization of Recombinant Cu,Zn Superoxide Dismutase fromPhotobacterium leiognathiExpressed inEscherichia coli:  Evidence for a Novel Catalytic Copper Binding Site

Biochemistry ◽  
1997 ◽  
Vol 36 (23) ◽  
pp. 7109-7113 ◽  
Author(s):  
Dolly Foti ◽  
Bruno Lo Curto ◽  
Giovanni Cuzzocrea ◽  
M. Elena Stroppolo ◽  
Francesca Polizio ◽  
...  
Keyword(s):  
Superoxide Dismutase ◽  
Binding Site ◽  
Spectroscopic Characterization ◽  
Copper Binding ◽  
Inescherichia Coli

scholarly journals Cloning and characterization of a haloarchaeal heat shock protein 70 functionally expressed inEscherichia coli

2007 ◽  
Vol 275 (1) ◽  
pp. 168-174 ◽  
Author(s):  
Hao Zhang ◽  
Lu Lin ◽  
Chi Zeng ◽  
Ping Shen ◽  
Yu-Ping Huang
Keyword(s):  
Heat Shock ◽  
Heat Shock Protein ◽  
Heat Shock Protein 70 ◽  
Inescherichia Coli
Sign in / Sign up

Export Citation Format

Share Document