Proteome-based identification of signal peptides for improved secretion of recombinant cyclomaltodextrin glucanotransferase in Escherichia coli

Ling, How Lie; Rahmat, Zaidah; Murad, Abdul Munir Abdul; Mahadi, Nor Muhammad; Illias, Rosli Md.

doi:10.1016/j.procbio.2017.06.018

Cited by 11 publications

(3 citation statements)

References 47 publications

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“…The results showed that the removal of SP remarkably enhanced the extracellular expression level of γ-CGTase. This was different from other extracellular-expressed CGTase described previously, ,, which indicated that the extracellular expression of recombinant CGTase is dependent on SPs, especially the Sec type. A suitable SP was believed to greatly improve the expression level.…”

Section: Discussioncontrasting

confidence: 99%

Regulation of Cell Membrane Permeability Enhanced the Non-Classical Secretion of γ-Cyclodextrin Glycosyltransferase in Bacillus subtilis

Jiang

Xie

et al. 2022

J. Agric. Food Chem.

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γ-Cyclodextrin glycosyltransferase EC 2.4.1.19) is an essential enzyme required in the production of γcyclodextrin, which shows huge prospects in the food, medicine, materials, and chemical industries. In this study, γ-CGTase from Bacillus sp. G-825-6 STB17 was successfully cloned and expressed in Bacillus subtilis WB600. The final extracellular activity of γ-CGTase can reach 45.34 U/mL with the deletion of the signal peptide, which was about 11.3 times of the initial level of γ-CGTase secreted by the general pathway. By monitoring the whole cultivation process, secretion was divided into two stages, which were dominated by cell membrane changes and apoptosis. The measurement of lactate dehydrogenase and the results of fluorescence microscopy demonstrated that the cell membrane permeability changed significantly in the middle stage of fermentation, proving that it played a crucial role in the non-classical secretion of γ-CGTase. Furthermore, the addition of Triton X-100, a non-ionic surfactant, remarkably enhanced the secretion level of γ-CGTase by 21.5%, which was caused by the increase in cell membrane permeability. This work is the first to obtain the extracellular expression of CGTase via the non-classical secretion pathway in B. subtilis and provides a new strategy to enhance extracellular expression by regulating the cell membranes.

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Section: Discussioncontrasting

confidence: 99%

Regulation of Cell Membrane Permeability Enhanced the Non-Classical Secretion of γ-Cyclodextrin Glycosyltransferase in Bacillus subtilis

Jiang

Xie

et al. 2022

J. Agric. Food Chem.

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“…Many heterologous proteins from Bacillus spp. are secreted to the extracellular space upon overexpression in E. coli , so evaluating the Bacillus secretome can yield new secretion signals [104]. Illias and colleagues computationally predicted signal sequences for 87 proteins highly secreted by Bacillus lehensis .…”

Section: Two-step Systemsmentioning

confidence: 99%

Developing Gram-negative bacteria for the secretion of heterologous proteins

et al. 2018

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Gram-negative bacteria are attractive hosts for recombinant protein production because they are fast growing, easy to manipulate, and genetically stable in large cultures. However, the utility of these microbes would expand if they also could secrete the product at commercial scales. Secretion of biotechnologically relevant proteins into the extracellular medium increases product purity from cell culture, decreases downstream processing requirements, and reduces overall cost. Thus, researchers are devoting significant attention to engineering Gram-negative bacteria to secrete recombinant proteins to the extracellular medium. Secretion from these bacteria operates through highly specialized systems, which are able to translocate proteins from the cytosol to the extracellular medium in either one or two steps. Building on past successes, researchers continue to increase the secretion efficiency and titer through these systems in an effort to make them viable for industrial production. Efforts include modifying the secretion tags required for recombinant protein secretion, developing methods to screen or select rapidly for clones with higher titer or efficiency, and improving reliability and robustness of high titer secretion through genetic manipulations. An additional focus is the expression of secretion machineries from pathogenic bacteria in the “workhorse” of biotechnology, Escherichia coli, to reduce handling of pathogenic strains. This review will cover recent advances toward the development of high-expressing, high-secreting Gram-negative production strains.

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“…Extracellular proteins of B. lehensis were subjected to 2-DE analysis, producing an extracellular proteome map [1] . A total of 87 identified proteins on the 2-DE was listed in Table 1 .…”

Section: Datamentioning

confidence: 99%

Data for proteome analysis of Bacillus lehensis G1 in starch-containing medium

et al. 2017

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Bacillus lehensis G1 is a cyclodextrin glucanotransferase (CGTase) producer, which can degrade starch into cyclodextrin. Here, we present the proteomics data of B. lehensis cultured in starch-containing medium, which is related to the article “Proteome-based identification of signal peptides for improved secretion of recombinant cyclomaltodextrin glucanotransferase in Escherichia coli” (Ling et. al, in press). This dataset was generated to better understand the secretion of proteins involved in starch utilization for bacterial sustained growth. A 2-DE proteomic technique was used and the proteins were tryptically digested followed by detection using MALDI-TOF/TOF. Proteins were classified into functional groups using the information available in SubtiList webserver (http://genolist.pasteur.fr/SubtiList/).

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Proteome-based identification of signal peptides for improved secretion of recombinant cyclomaltodextrin glucanotransferase in Escherichia coli

Cited by 11 publications

References 47 publications

Regulation of Cell Membrane Permeability Enhanced the Non-Classical Secretion of γ-Cyclodextrin Glycosyltransferase in Bacillus subtilis

Regulation of Cell Membrane Permeability Enhanced the Non-Classical Secretion of γ-Cyclodextrin Glycosyltransferase in Bacillus subtilis

Developing Gram-negative bacteria for the secretion of heterologous proteins

Data for proteome analysis of Bacillus lehensis G1 in starch-containing medium

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