High-level production of recombinant sulfide-reactive hemoglobin I from Lucina pectinata in Escherichia coliHigh yields of fully functional holoprotein synthesis in the BLi5 E. coli strain

2004 ◽  
Vol 38 (2) ◽  
pp. 184-195 ◽  
Author(s):  
R LEON ◽  
H MUNIERLEHMANN ◽  
O BARZU ◽  
V BAUDINCREUZA ◽  
R PIETRI ◽  
...  
2015 ◽  
Vol 14 (1) ◽  
Author(s):  
Zhe Zhang ◽  
Grietje Kuipers ◽  
Łukasz Niemiec ◽  
Thomas Baumgarten ◽  
Dirk Jan Slotboom ◽  
...  

2012 ◽  
Vol 47 (3) ◽  
pp. 395-400 ◽  
Author(s):  
Zhong Yang ◽  
Rongrong Ma ◽  
Lei Huang ◽  
Xiangcheng Zhu ◽  
Jiayuan Sheng ◽  
...  

2011 ◽  
Vol 66 (11-12) ◽  
pp. 605-613 ◽  
Author(s):  
He Wang ◽  
Ruijin Yang ◽  
Xiao Hua ◽  
Zhong Zhang ◽  
Wei Zhao ◽  
...  

High-level production of recombinant glucose isomerase (rGI) is desirable for lactulose synthesis. In this study, the xylA gene encoding glucose isomerase from Actinoplanes missouriensis CICIM B0118(A) was cloned and expressed in E. coli BL21(DE3), and high-level production was performed by optimization of the medium composition. rGI was purified from a recombinant E. coli BL21(DE3) and characterized. The optimum pH value of the purified enzyme was 8.0 and it was relatively stable within the pH range of 7.0 - 9.0. Its optimum temperature was around 85 °C, and it exhibited good thermostability when the temperature was lower than 90 °C. The maximum enzyme activity required the presence of both Co2+ and Mg2+, at the concentrations of 200 μM and 8 mM, respectively. With high-level expression and the simple one-step chromatographic purification of the His-tagged recombinant enzyme, this GI could be used in industrial production of lactulose as a potential economic tool


2004 ◽  
Vol 38 (2) ◽  
pp. 184-195 ◽  
Author(s):  
Ruth Gretchen León ◽  
Hélène Munier-Lehmann ◽  
Octavian Barzu ◽  
Véronique Baudin-Creuza ◽  
Ruth Pietri ◽  
...  

2009 ◽  
Vol 77 (6) ◽  
pp. 2272-2284 ◽  
Author(s):  
Chantal Peigne ◽  
Philippe Bidet ◽  
Farah Mahjoub-Messai ◽  
Céline Plainvert ◽  
Valérie Barbe ◽  
...  

ABSTRACT A new Escherichia coli virulent clonal group, O45:K1, belonging to the highly virulent subgroup B21 was recently identified in France, where it accounts for one-third of E. coli neonatal meningitis cases. Here we describe the sequence, epidemiology and function of the large plasmid harbored by strain S88, which is representative of the O45:K1 clonal group. Plasmid pS88 is 133,853 bp long and contains 144 protein-coding genes. It harbors three different iron uptake systems (aerobactin, salmochelin, and the sitABCD genes) and other putative virulence genes (iss, etsABC, ompT P, and hlyF). The pS88 sequence is composed of several gene blocks homologous to avian pathogenic E. coli plasmids pAPEC-O2-ColV and pAPEC-O1-ColBM. PCR amplification of 11 open reading frames scattered throughout the plasmid was used to investigate the distribution of pS88 and showed that a pS88-like plasmid is present in other meningitis clonal groups such as O18:K1, O1:K1, and O83:K1. A pS88-like plasmid was also found in avian pathogenic strains and human urosepsis strains belonging to subgroup B21. A variant of S88 cured of its plasmid displayed a marked loss of virulence relative to the wild-type strain in a neonatal rat model, with bacteremia more than 2 log CFU/ml lower. The salmochelin siderophore, a known meningovirulence factor, could not alone explain the plasmid's contribution to virulence, as a salmochelin mutant displayed only a minor fall in bacteremia (0.9 log CFU/ml). Thus, pS88 is a major virulence determinant related to avian pathogenic plasmids that has spread not only through meningitis clonal groups but also human urosepsis and avian pathogenic strains.


2017 ◽  
Vol 15 (4) ◽  
pp. 4-12
Author(s):  
Elena A. Dolgikh ◽  
Anna N. Kirienko ◽  
Oksana D. Kovaleva ◽  
Igor A. Tikhonovich

Background. Rhizobial Nod factors (NFs), the key regulators of legume-rhizobia symbiosis, act in low concentrations and their biological activity depends on structural features, that suggests the presence of specific receptors in plants. Putative receptors, LysM-receptor-like kinases (LysM-RLKs), were found in model legumes L. japonicus and M. truncatula. However, binding capacity with NFs was only studied for L. japonicus LysM-RLKs. In pea a few candidates for NF receptors like Sym10, Sym37 and K1 were found. Analysis of mutants revealed the importance of these proteins for symbiosis development. However, the biochemical function of these receptors has not been studied. Materials and methods. Sequences encoding extracellular domains (ECDs) of LysM-RLKs Sym10, Sym37, and K1 were cloned in the pRSETa vector. Constructs were introduced in E. coli strain C41 to produce proteins with His6 residues on either the amino or carboxyl terminus. Protein purification was carried out using metal chelate affinity chromatography. The binding capacity with ligand was evaluated using ProteonXPR36 biosensor. Results. To study binding capacity with NFs, we have developed approaches for the synthesis of LysM-RLK Sym10, Sym37 and K1 in soluble form in heterologous system. The high level of protein synthesis was achieved at +28 °C using 0,5 mM IPTG in 2-16 hours. Analysis of binding capacity of ECDs with NFs revealed the low affinity using the surface plasmon resonance. Conclusion. The possibility of recombinant receptor synthesis in soluble state in E. coli at high level was demonstrated. Analysis of binding capacity with NFs showed the potential interaction, but with low affinity.


2004 ◽  
Vol 26 (19) ◽  
pp. 1501-1504 ◽  
Author(s):  
Seung-Hwan Jang ◽  
Dong-Ku Kang ◽  
Soo-Ik Chang ◽  
Harold A. Scheraga ◽  
Hang-Cheol Shin
Keyword(s):  
E Coli ◽  

2019 ◽  
Author(s):  
Ching Hei Phoebe Cheung ◽  
Jacqueline Findlay ◽  
Kate J. Heesom ◽  
Matthew B. Avison

AbstractBackgroundBicyclic boronates are a new and potentially important class of β-lactamase inhibitor, with the ability to inhibit β-lactamases from all molecular classes, including mobile metallo-β-lactamases.ObjectiveOur objective was to identify mutants resistant to the actions of the bicyclic boronate inhibitor 2, when being used in combination with aztreonam.MethodsOvernight cultures were plated on to agar containing increasing concentrations of aztreonam with a fixed 10 mg/L concentration of the inhibitor. Resistant derivatives and parent strains were analysed by whole genome sequencing and LC-MS/MS proteomics to identify mechanism of resistance.ResultsWhen using a mixed overnight culture containing one Escherichia coli (TEM-1, CTX-M-15, CMY-4 producer) and one Klebsiella pneumoniae (SHV-12, CTX-M-15, NDM-1 producer) mobilisation of an IncX3 plasmid carrying blaSHV-12 from the K. pneumoniae into the E. coli generated an aztreonam/boronate resistant derivative.ConclusionsHigh-level production of three bicyclic boronate susceptible enzymes (CMY-4, CTX-M-15, SHV-12) capable of hydrolysing aztreonam plus TEM-1, which binds the inhibitor, overcomes the fixed inhibitor dose used. This was only identified when using a mixed culture for selection. It would seem prudent that to allow for coalescence of the myriad β-lactamase genes commonly found in bacterial populations colonising humans, this mixed culture approach should be the norm when testing the potential for generating β-lactamase inhibitor resistance in pre-clinical analysis.


Sign in / Sign up

Export Citation Format

Share Document