RESUMO
Any unmanned surface ship is subject to system uncertainty, unknown parameters, and external disturbance induced by the wind, the wave loads, and the ocean currents. They may deteriorate ship's control accuracy. This paper aims to solve the trajectory tracking control problem of unmanned surface ships with disturbance and system uncertainty accommodated simultaneously. An estimator-based backstepping controller is presented with an estimator designed to provide a precise estimation of the disturbance and uncertainties. The proposed controller ensures the closed-loop tracking system to be globally exponentially stable. The trajectory tracking error and the estimation error of disturbance and uncertainties are globally exponentially stable. The key feature of the developed control scheme is that it is more robust to disturbances and system uncertainties. Simulation results are further presented to validate the effectiveness of the approach.
RESUMO
Hyaluronic acid (HA), a vital acid mucopolysaccharide, has immense applied value in foodstuffs, medicaments, and cosmetics among others. UDP-glucose dehydrogenase (UGDH, EC 1.1.1.22) is an essential enzyme for HA synthesis. In this study, a UGDH (PmuHasB, 45.9 kDa) from Pasteurella multocida CVCC 408 was expressed in Escherichia coli BL21 (DE3). It was purified by two chromatographic columns with a specific activity of 6.58 IU/mg. The optimum pH and temperature were determined to be 10.0 and 37°C, respectively. The activity was stable across the pH range 6-10, and had a half-life of about 3 h at 45°C. The estimated apparent Km values for UDP-glucose and NAD(+) were 0.11 and 0.069 mM, respectively. The results indicated that PmuHasB was an alkaline and mesophilic UGDH. PmuHasB and PmuHasA (HA synthase, HAS) were co-expressed in E. coli BW25113 to obtain a HA high-producing strain pBPAB/BW25113. It produced about 2.39 g/L HA in shake flask by using the method of whole-cell catalysis. Investigation of the different UGDHs on HA synthesis revealed that intracellular UGDH activity and HA total yield of pBPAB/BW25113 (0.15 IU/mg and 5.4 g/L) were higher than from pBPASB/BW25113 (0.013 IU/mg and 2.8 g/L) and pBPAEB/BW25113 (0.010 IU/mg and 2.22 g/L). These results indicated that the activity and stability of UGDH plays a significant role in HA production, and should prove useful for further genetic engineering research with a view to construct other glucuronic acid polysaccharide synthesis pathways.
Assuntos
Proteínas de Bactérias/metabolismo , Ácido Hialurônico/biossíntese , Pasteurella multocida/enzimologia , Uridina Difosfato Glucose Desidrogenase/metabolismo , Proteínas de Bactérias/química , Proteínas de Bactérias/genética , Estabilidade Enzimática , Escherichia coli/genética , Genes Bacterianos , Cinética , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Uridina Difosfato Glucose Desidrogenase/química , Uridina Difosfato Glucose Desidrogenase/genéticaRESUMO
We amplified genes encoding UDP-glucose dehydrogenase, ecohasB from Escherichia coli and spyhasB from Streptococcus pyogenes. Both ecohasB and spyhasB were inserted into T7 expression vector pRX2 to construct recombinant plasmids pRXEB and pRXSB, and to express in E. coli BL21(DE3). After nickel column purification of UDP-glucose dehydrogenases, the enzymes were characterized. The optimum reaction condition of spyHasB was at 30 °C and pH 10. The specific activity reached 12.2 U/mg under optimum condition. The optimum reaction condition of ecoHasB was at 30 °C and pH 9. Its specific activity reached 5.55 U/mg under optimum condition. The pmuhasA gene encoding hyaluronic acid synthase was amplified from Pasteurella multocida and ligated with ecohasB and spyhasB to construct the coexpression vectors pBPAEB and pBPASB, respectively. The co-expression vectors were transformed into E. coli BW25113. Hyaluronic acid (HA) was produced by biotransformation and the conditions were optimized. When recombinant strains were used to produce hyaluronic acid, the higher the activity of UDP-glucose dehydrogenase was, the better its stability was, and the higher the HA production could reach. Under the optimal conditions, the yields of HA produced by pBPAEB/BW25113 and pBPASB/BW25113 in shake flasks were 1.52 and 1.70 g/L, respectively, and the production increased more than 2-3 folds as previously reported.
