Cloning and Sequence Analysis of Mutant OmpF from Antibiotic Resistant Escherichia coli

[Objective]This study aimed to investigate the mutations of OmpF from an isolated antibiotic resistant Escherichia coli strain.[Methods]The mutant OmpF（mOmpF）from antibiotic resistant E.coli was amplified by PCR with Pfu DNA polymerase and ligated into the expression vector pET28a.Subsequently,the expression vector pET28-mOmpF was sequenced and analyzed by DNAMAN software and Swiss-Model online.[Result]Sequence analysis revealed that the open reading fragment of mOmpF was 903 bp long,which was mutated dramatically compared to that of the 1 020 bp long model OmpF.The DNA sequence shared only54.5%homology with OmpF.mOmpF was 44.6%identical to that of OmpF.Protein structure predication and analysis through Swiss-Model online suggested that the structure of mOmpF changed dramatically compared to OmpF.[Conclusion]The present study provided basis for further analyzing the relationships between the structure and functions of mOmpF from antibiotic resistant E.coli.

Amino terminus mutant OmpA from an isolated antibiotic resistant <i>Escherichia</i><i>coli</i>still possess resistance to environmental stresses

Antibiotic resistant Escherichia coli strains are becoming more common recently. OmpA is a very important antigen protein of E. coli, which consists of two separate domains, N-terminal and C-terminal domain. The N-terminal domain contains eight β- barrel regions that plays important roles in the multifaceted functions of OmpA. In the present study, we cloned a mutant OmpA gene from a multi-antibiotic resistant E. coli strain. Sequence analysis indicated that the N-terminal DNA sequence of the mutant OmpA shared 81.05% homology with the modeled OmpA from E. coli K12 and the N-terminal amino acid sequence of the mutant OmpA was 81.22% identical to that of the E. coli K12 OmpA. Moreover, several amino acids located in the β-barrel region were mutated. The mutant OmpA was expressed in BL21 suggested by SDS-PAGE. Resistance to environmental stress assay indicated that the N-terminus mutant OmpA still possessed excellent activities in pH, temperature and osmotic pressure resistance. Our pre- sent study may supply insights into better and deeper understand the relationships between OmpA N-terminal regions and its functions in environmental stress conditions and the mechanisms on antibiotic resistance of E. coli.

Considering for the blanket structure scheme of HCCB DEMO

For the solid blanket concept of helium cooled ceramic breeder (HCCB) demonstration fusion power plant (DEMO), a feasible blanket structure with configuration 2×X is proposed as considering relatively low temperature limit of neutron multiplier beryllium pebbles. Based on that, preliminary design for the typical blanket module of HCCB DEMO has been carried out and verified by thermal-hydraulic analysis and structural analysis. Furthermore, the specific relationship of maximum temperature depended on the surface heating of blanket key part first wall (FW) is also analyzed.

Spectral properties of LH2 exhibit very similar even when heterologously express LH2 with <i>β</i>-subunit fusion protein in <i>Rhodobacter</i><i>sphaeroides</i>

Interactions between the light-harvesting subunits and the non-covalently bound photopigments attribute considerably to the spectral properties of photosynthetic bacteria light-harvesting complexes. In our previous studies, we have constructed a novel Rhodobacter sphaeroides expression system. In the present study, we focus on the spectral properties of LH2 when heterologously express LH2 with β-subunit- GFP fusion protein in Rb. sphaeroides. Near infra-red spectrum of LH2 remained nearly unchanged as measured by spectroscopy. Fluorescence spectrum suggested that the LH2 with β-subunit-GFP fusion protein complexes still possessed normal activity in energy transfer. However, photopigments contents were significantly decreased to a very low level in the LH2 with β-subunit-GFP fusion protein complexes compared to that of LH2. FT-IR spectra indicated that interactions between photopigments and LH2 α/β- subunits appeared not to be changed. It was concluded that the LH2 spectral properties exhibited very similar even when heterologously expressed LH2 b-subunit fusion protein in Rb. sphaeroides. Our present study may supply a new insight into better understand the interactions between light-harvesting subunits and photopigments and bacterial photosynthesis and promote the development of the novel Rb. sphaeroides expression system.