scholarly journals Improving the production of AHL lactonase AiiO-AIO6 from Ochrobactrum sp. M231 in intracellular protease-deficient Bacillus subtilis

2020 ◽  
Author(s):  
Rui Xia ◽  
Yalin Yang ◽  
Xingliang Pan ◽  
Chenchen Gao ◽  
Yuanyuan Yao ◽  
...  
Keyword(s):  
Bacillus Subtilis ◽  
Positive Impact ◽  
Cell Communication ◽  
Quorum Quenching ◽  
Catalytic Triad ◽  
Heterologous Proteins ◽  
Extracellular Production ◽  
Secretion Pathway ◽  
Protease Deficient ◽  
Encoding Genes

Abstract Quorum quenching (QQ) blocks bacterial cell-to-cell communication (i.e., quorum sensing), and is a promising antipathogenic strategy to control bacterial infection via inhibition of virulence factor expression and biofilm formation. QQ enzyme AiiO-AIO6 from Ochrobactrum sp. M231 has several excellent properties and shows biotherapeutic potential against important bacterial pathogens of aquatic species. AiiO-AIO6 can be secretory expressed in Bacillus subtilis via a non-classical secretion pathway. To improve AiiO-AIO6 production, four intracellular protease-deletion mutants of B. subtilis 1A751 were constructed by individually knocking out the intracellular protease-encoding genes ( tepA, ymfH, yrrN and ywpE ). The AiiO-AIO6 expression plasmid pWB-AIO6BS was transformed into the B. subtilis 1A751 and its four intracellular protease-deletion derivatives. Results showed that all recombinant intracellular protease-deletion derivatives (BSΔ tepA , BSΔ ymfH , BSΔ yrrN and BSΔ ywpE ) had a positive impact on AiiO-AIO6 production. The highest amount of AiiO-AIO6 extracellular production of BSΔ ywpE in shake flask reached 3530 U/mL, which was about 62% higher than that of the wild-type strain. Furthermore, LC-MS/MS analysis of the degrading products of 3-oxo-C8-HSL by purification of AiiO-AIO6 indicated that AiiO-AIO6 was an AHL-lactonase which hydrolyzes the lactone ring of AHLs. Phylogenetic analysis showed that AiiO-AIO6 was classified as a member of the α/β hydrolase family with a conserved “nucleophile-acid-histidine” catalytic triad. In summary, this study showed that intracellular proteases were responsible for the reduced yields of heterologous proteins and provided an efficient strategy to enhance the extracellular production of AHL lactonase AiiO-AIO6.

scholarly journals Improving the production of AHL lactonase AiiO-AIO6 from Ochrobactrum sp. M231 in intracellular protease-deficient Bacillus subtilis

2020 ◽  
Author(s):  
Rui Xia ◽  
Yalin Yang ◽  
Xingliang Pan ◽  
Chenchen Gao ◽  
Yuanyuan Yao ◽  
...  
Keyword(s):  
Bacillus Subtilis ◽  
Positive Impact ◽  
Cell Communication ◽  
Quorum Quenching ◽  
Catalytic Triad ◽  
Heterologous Proteins ◽  
Extracellular Production ◽  
Secretion Pathway ◽  
Protease Deficient ◽  
Encoding Genes

Abstract Quorum quenching (QQ) blocks bacterial cell-to-cell communication (i.e., quorum sensing), and is a promising antipathogenic strategy to control bacterial infection via inhibition of virulence factor expression and biofilm formation. QQ enzyme AiiO-AIO6 from Ochrobactrum sp. M231 has several excellent properties and shows biotherapeutic potential against important bacterial pathogens of aquatic species. AiiO-AIO6 can be secretory expressed in Bacillus subtilis via a non-classical secretion pathway. To improve AiiO-AIO6 production, four intracellular protease-deletion mutants of B. subtilis 1A751 were constructed by individually knocking out the intracellular protease-encoding genes (tepA, ymfH, yrrN and ywpE). The AiiO-AIO6 expression plasmid pWB-AIO6BS was transformed into the B. subtilis 1A751 and its four intracellular protease-deletion derivatives. Results showed that all recombinant intracellular protease-deletion derivatives (BSΔtepA, BSΔymfH, BSΔyrrN and BSΔywpE) had a positive impact on AiiO-AIO6 production. The highest amount of AiiO-AIO6 extracellular production of BSΔywpE in shake flask reached 3530 U/mL, which was about 62% higher than that of the wild-type strain. Furthermore, LC-MS/MS analysis of the degrading products of 3-oxo-C8-HSL by purification of AiiO-AIO6 indicated that AiiO-AIO6 was an AHL-lactonase which hydrolyzes the lactone ring of AHLs. Phylogenetic analysis showed that AiiO-AIO6 was classified as a member of the α/β hydrolase family with a conserved “nucleophile-acid-histidine” catalytic triad. In summary, this study showed that intracellular proteases were responsible for the reduced yields of heterologous proteins and provided an efficient strategy to enhance the extracellular production of AHL lactonase AiiO-AIO6.

scholarly journals Improving the production of AHL lactonase AiiO-AIO6 from Ochrobactrum sp. M231 in intracellular protease-deficientBacillus subtilis

2020 ◽  
Author(s):  
Rui Xia ◽  
Yalin Yang ◽  
Xingliang Pan ◽  
Chenchen Gao ◽  
Yuanyuan Yao ◽  
...  
Keyword(s):  
Wild Type Strain ◽  
Positive Impact ◽  
Cell Communication ◽  
Quorum Quenching ◽  
Catalytic Triad ◽  
Heterologous Proteins ◽  
Extracellular Production ◽  
Expression Plasmid ◽  
Encoding Genes ◽  
Hydrolase Family

Abstract Quorum quenching (QQ) blocks bacterial cell-to-cell communication (i.e., quorum sensing), and is a promising antipathogenic strategy to control bacterial infection via inhibition of virulence factor expression and biofilm formation. QQ enzyme AiiO-AIO6 from Ochrobactrum sp. M231 has several excellent properties and shows biotherapeutic potential against important bacterial pathogens of aquatic species. AiiO-AIO6 can be secretory expressed in Bacillus subtilis via a non-classical secretion pathway.To improve AiiO-AIO6 production, four intracellular protease-deletion mutants of B. subtilis1A751 were constructed by individually knocking out the intracellular protease-encoding genes (tepA, ymfH, yrrNandywpE). The AiiO-AIO6 expression plasmid pWB-AIO6BS was transformed into the B. subtilis 1A751 and its four intracellular protease-deletion derivatives. Results showed that all recombinant intracellular protease-deletion derivatives (BSΔtepA, BSΔymfH, BSΔyrrNand BSΔywpE) had a positive impact on AiiO-AIO6 production. The highest amount of AiiO-AIO6 extracellular production of BSΔywpE in shake flask reached 1416.47 U/mL/OD600, which was about 121% higher than that of the wild-type strain. Furthermore, LC-MS/MS analysis of the degrading products of 3-oxo-C8-HSL by purification of AiiO-AIO6 indicated that AiiO-AIO6 was an AHL-lactonase which hydrolyzes the lactone ring of AHLs. Phylogenetic analysis showed that AiiO-AIO6was classified as a member of the α/β hydrolase family with a conserved “nucleophile-acid-histidine” catalytic triad. In summary, this study showed that intracellular proteases were responsible for the reduced yields of heterologous proteins and provided an efficient strategy to enhance the extracellular production of AHL lactonase AiiO-AIO6.

Enhanced Extracellular Production of Heterologous Proteins in Bacillus subtilis by Deleting the C-terminal Region of the SecA Secretory Machinery

2010 ◽  
Vol 46 (3) ◽  
pp. 250-257 ◽  
Author(s):  
Hiroshi Kakeshtia ◽  
Yasushi Kageyama ◽  
Katsutoshi Ara ◽  
Katsuya Ozaki ◽  
Kouji Nakamura
Keyword(s):  
Bacillus Subtilis ◽  
Terminal Region ◽  
Heterologous Proteins ◽  
Extracellular Production

Probiotics in the diets of laying quails

2020 ◽  
pp. 60-66
Author(s):  
O. Merzlyakova ◽  
V. Rogachyev ◽  
V. Chegodaev
Keyword(s):  
Bacillus Subtilis ◽  
Bacillus Licheniformis ◽  
Egg Production ◽  
Positive Impact ◽  
Hematological Parameters ◽  
Economic Effect ◽  
Egg Laying ◽  
Control Group ◽  
Egg Mass ◽  
The Cost

The efficiency of introducing probiotics based on strains of Bacillus subtilis, Bacillus licheniformis and their consortium in the amount of 150 g/t of feed into the diets of laying quails has been studied. The experiment lasting 182 days has been carried out on four groups of quails with 30 heads in each. The quails have been housed in the broiler battery in compliance with the required microclimate conditions. Quails of all groups have been received the main diet (compound feed) developed taking into account their age and physiological characteristics. The quails of the 1st, 2nd and 3rd experimental groups in addition to the main diet received probiotics (150 g/t compound feed) based on strains Bacillus subtilis, Bacillus licheniformis and their consortium, respectively. It has been found that feeding the laying quails of the consortium of strains Bacillus subtilis and Bacillus licheniformis had the most significant positive impact on their productive performance, it allowed to increase egg production by 7,81 %, egg laying intensity by 5,0 %, egg mass yield by 9,77 %, while reducing feed expenditures for 10 eggs by 13,35 %. The yield of hatching eggs has been increased by 7,03 %, hatchability of chickens from laid and fertilized eggs by 8,33 and 8,35 %, brooding waste decreased by 21,74 %. Hematological parameters of quails during the whole experiment were within the physiological norm. The economic effect calculated on the basis of data on the cost of compound feed, probiotics and the cost of sold eggs of quail laying was 14,56 % in the 3rd experimental group (in relation to the control group).

scholarly journals Substrate-Induced Production and Secretion of Cellulases by Clostridium acetobutylicum

2004 ◽  
Vol 70 (9) ◽  
pp. 5238-5243 ◽  
Author(s):  
Ana M. López-Contreras ◽  
Krisztina Gabor ◽  
Aernout A. Martens ◽  
Bernadet A. M. Renckens ◽  
Pieternel A. M. Claassen ◽  
...  
Keyword(s):  
Catabolite Repression ◽  
Clostridium Acetobutylicum ◽  
Glycoside Hydrolase ◽  
Protein A ◽  
Cellulase Activity ◽  
Glycoside Hydrolase Family ◽  
Extracellular Production ◽  
Encoding Genes ◽  
Glycoside Hydrolase Family 48 ◽  
Hydrolase Family

ABSTRACT Clostridium acetobutylicum ATCC 824 is a solventogenic bacterium that grows heterotrophically on a variety of carbohydrates, including glucose, cellobiose, xylose, and lichenan, a linear polymer of β-1,3- and β-1,4-linked β-d-glucose units. C. acetobutylicum does not degrade cellulose, although its genome sequence contains several cellulase-encoding genes and a complete cellulosome cluster of cellulosome genes. In the present study, we demonstrate that a low but significant level of induction of cellulase activity occurs during growth on xylose or lichenan. The celF gene, located in the cellulosome-like gene cluster and coding for a unique cellulase that belongs to glycoside hydrolase family 48, was cloned in Escherichia coli, and antibodies were raised against the overproduced CelF protein. A Western blot analysis suggested a possible catabolite repression by glucose or cellobiose and an up-regulation by lichenan or xylose of the extracellular production of CelF by C. acetobutylicum. Possible reasons for the apparent inability of C. acetobutylicum to degrade cellulose are discussed.

scholarly journals Complete Genome Sequence of Bacillus subtilis Strain WB800N, an Extracellular Protease-Deficient Derivative of Strain 168

2018 ◽  
Vol 7 (18) ◽  
Author(s):  
Haeyoung Jeong ◽  
Da-Eun Jeong ◽  
Seung-Hwan Park ◽  
Seong Joo Kim ◽  
Soo-Keun Choi
Keyword(s):  
Bacillus Subtilis ◽  
Genome Sequence ◽  
Complete Genome Sequence ◽  
Complete Genome ◽  
Genetically Engineered ◽  
Subtilis Strain ◽  
Secretory Proteins ◽  
Extracellular Proteases ◽  
Content Type ◽  
Protease Deficient

Bacillus subtilis WB800N is a genetically engineered variant of B. subtilis 168, such that all extracellular proteases are disrupted, which enables WB800N to be widely used for the expression of secretory proteins. Here, we report the 4.2-Mb complete genome sequence of WB800N and present all of the disrupted gene structure.

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