Export of recombinant proteins in Escherichia coli using ABC transporter with an attached lipase ABC transporter recognition domain (LARD)

<p>Abstract</p> <p>Background</p> <p>ATP binding cassette (ABC) transporter secretes the protein through inner and outer membranes simultaneously in gram negative bacteria. Thermostable lipase (TliA) of <it>Pseudomonas fluorescens </it>SIK W1 is secreted through the ABC transporter. TliA has four glycine-rich repeats (GGXGXD) in its C-terminus, which appear in many ABC transporter-secreted proteins. From a homology model of TliA derived from the structure of <it>P. aeruginosa </it>alkaline protease (AprA), lipase ABC transporter domains (LARDs) were designed for the secretion of fusion proteins.</p> <p>Results</p> <p>The LARDs included four glycine-rich repeats comprising a β-roll structure, and were added to the C-terminus of test proteins. Either Pro-Gly linker or Factor Xa site was added between fusion proteins and LARDs. We attached different length of LARDs such as LARD0, LARD1 or whole TliA (the longest LARD) to three types of proteins; green fluorescent protein (GFP), epidermal growth factor (EGF) and cytoplasmic transduction peptide (CTP). These fusion proteins were expressed in <it>Escherichia coli </it>together with ABC transporter of either <it>P. fluorescens </it>or <it>Erwinia chrysanthemi</it>. Export of fusion proteins with the whole TliA through the ABC transporter was evident on the basis of lipase enzymatic activity. Upon supplementation of <it>E. coli </it>with ABC transporter, GFP-LARDs and EGF-LARDs were excreted into the culture supernatant.</p> <p>Conclusion</p> <p>The LARDs or whole TliA were attached to C-termini of model proteins and enabled the export of the model proteins such as GFP and EGF in <it>E. coli </it>supplemented with ABC transporter. These results open the possibility for the extracellular production of recombinant proteins in <it>Pseudomonas </it>using LARDs or TliA as a C-terminal signal sequence.</p

VALIDATION OF OPTIMALLY DESIGNED STATOR-PROPELLER SYSTEM BY EFD AND CFD

The development of energy-saving devices to lower the energy efficiency design index (EEDI) of ships has been actively researched worldwide. One such device is an asymmetric pre-swirl stator, which helps to improve the propulsion efficiency by recovering the rotational energy generated during propeller rotation. Determining the pitch angle is the most important factor in the design of an efficient asymmetric pre-swirl stator. To optimize the pitch angle of an asymmetric pre-swirl stator, this study deals with potential-flow, computational fluid dynamics, and model tests. The model delivered power at a design speed of 24 kt was compared by changing the pitch angle by ±2° with respect to the reference angle designed using a potential-flow program. The commercial code Star-CCM+ was used for the numerical analysis, and the model was also tested in a towing tank at Pusan National University. This study proposes an effective method for determining and verifying the optimal pitch angle of an asymmetric pre-swirl stator

FEASIBILITY STUDY ON EFFECT OF STRUCTURAL FLEXIBILITY OF ASYMMETRIC PRE-SWIRL STATOR ON PROPULSION PERFORMANCE FOR KRISO CONTAINER SHIP (KCS)

The use of energy-saving devices is the most effective method for decreasing CO2 emissions, which is an increasingly concerning environmental issue. The asymmetric pre-swirl stator has been developed as an energy-saving device and has been successfully applied to various types of vessels. In the present study, a flexible material was applied to an asymmetric pre-swirl stator to determine the variation in the flow around stator and its efficiency. A fluid–structure interaction (FSI) analysis system was developed using the Star-CCM+ (fluid) and the Abaqus (structure). The proposed analysis system was validated by comparing the experimental results using a flexible plate in a flowing fluid. The flexible stator was applied to a 3,600 TEU KRISO Container Ship to determine the improvement in its performance compared to the previous optimum value achieved with a rigid stator. Although this application was conducted on a model scale and the deformation was small, the results of the flexible stator indicated the possibility of not only increasing the efficiency but also decreasing the vortex risk around stator blade