Enhanced production of heterologous proteins via engineering the cell surface of Bacillus licheniformis.

Publisher: Oxford University Press Country of Publication: Germany NLM ID: 9705544 Publication Model: Print-Electronic Cited Medium: Internet ISSN: 1476-5535 (Electronic) Linking ISSN: 13675435 NLM ISO Abbreviation: J Ind Microbiol Biotechnol Subsets: MEDLINE

Publication: 2021- : [Oxford] : Oxford University Press
Original Publication: Houndmills, Basingstoke, Hampshire, UK : Published by Stockton Press on behalf of the Society for Industrial Microbiology, c1996-

Bacillus licheniformis/*enzymology; Bacillus licheniformis/genetics ; Cell Membrane/enzymology ; Membrane Proteins/genetics ; Membrane Proteins/metabolism ; Protein Engineering ; Subtilisins/genetics ; Subtilisins/metabolism ; Transferases (Other Substituted Phosphate Groups)/genetics ; Transferases (Other Substituted Phosphate Groups)/metabolism ; alpha-Amylases/genetics ; alpha-Amylases/metabolism

Cell surface engineering was proven as the efficient strategy for enhanced production of target metabolites. In this study, we want to improve the yield of target protein by engineering cell surface in Bacillus licheniformis. First, our results confirmed that deletions of D-alanyl-lipoteichoic acid synthetase gene dltD, cardiolipin synthase gene clsA and CDP-diacylglycerol-serine O-phosphatidyltransferase gene pssA were not conducive to cell growth, and the biomass of gene deletion strains were, respectively, decreased by 10.54 ± 1.43%, 14.17 ± 1.51%, and 17.55 ± 1.28%, while the concentrations of total extracellular proteins were improved, due to the increases of cell surface net negative charge and cell membrane permeability. In addition, the activities of target proteins, nattokinase, and α-amylase were also improved significantly in gene deletion strains. Furthermore, the triplicate gene (dltD, clsA, and pssA) deletion strain was constructed, which further led to the 45.71 ± 2.43% increase of cell surface net negative charge and 26.45 ± 2.31% increase of cell membrane permeability, and the activities of nattokinase and α-amylase reached 37.15 ± 0.89 FU/mL and 305.3 ± 8.4 U/mL, increased by 46.09 ± 3.51% and 96.34 ± 7.24%, respectively. Taken together, our results confirmed that cell surface engineering via deleting dltD, clsA, and pssA is an efficient strategy for enhanced production of target proteins, and this research provided a promising host strain of B. licheniformis for efficient protein expression.

J Ind Microbiol Biotechnol. 2014 Nov;41(11):1599-607. (PMID: 25212246)
Int J Biol Macromol. 1994 Oct;16(5):265-75. (PMID: 7534473)
Appl Biochem Biotechnol. 2019 Apr;187(4):1502-1514. (PMID: 30267286)
Appl Microbiol Biotechnol. 2017 Feb;101(3):933-949. (PMID: 28062973)
J Bacteriol. 2001 Mar;183(6):2051-8. (PMID: 11222605)
J Ind Microbiol Biotechnol. 2015 Feb;42(2):287-95. (PMID: 25475755)
Microbiology. 2007 Jul;153(Pt 7):2126-36. (PMID: 17600057)
Lett Appl Microbiol. 2003;37(1):75-80. (PMID: 12803561)
J Biol Chem. 2000 Sep 1;275(35):26696-703. (PMID: 10871614)
Microb Cell Fact. 2017 Feb 8;16(1):24. (PMID: 28178978)
Front Microbiol. 2019 Feb 01;10:105. (PMID: 30774627)
Microb Cell Fact. 2015 Jun 26;14:92. (PMID: 26112883)
J Bacteriol. 2006 Aug;188(16):5797-805. (PMID: 16885447)
J Appl Microbiol. 2019 Jun;126(6):1632-1642. (PMID: 30609144)
J Appl Microbiol. 2005;99(2):363-75. (PMID: 16033468)
J Bacteriol. 2016 May 13;198(11):1585-1594. (PMID: 27002131)
Trends Microbiol. 2008 Feb;16(2):73-9. (PMID: 18182292)
Protein Expr Purif. 2018 Feb;142:81-87. (PMID: 28963005)
PLoS One. 2013;8(1):e54031. (PMID: 23349781)
ACS Synth Biol. 2017 Jun 16;6(6):1065-1075. (PMID: 28252945)
Biotechnol Bioeng. 2018 Oct;115(10):2541-2553. (PMID: 29940069)
Chem Rev. 2018 Jan 10;118(1):4-72. (PMID: 28443658)
Biotechnol Biofuels. 2018 Nov 9;11:306. (PMID: 30455735)
Biosci Biotechnol Biochem. 2016 Jul;80(7):1243-53. (PMID: 27305282)
J Biol Chem. 2004 Apr 30;279(18):19302-14. (PMID: 14976191)
Microb Cell Fact. 2017 Apr 24;16(1):70. (PMID: 28438200)
Microb Cell Fact. 2017 Apr 4;16(1):56. (PMID: 28376879)
Microb Cell Fact. 2017 Apr 4;16(1):57. (PMID: 28376795)
Appl Microbiol Biotechnol. 2017 May;101(10):4151-4161. (PMID: 28197687)
Sci Rep. 2017 Feb 23;7:43404. (PMID: 28230096)
Biotechnol Bioeng. 2018 Jan;115(1):216-231. (PMID: 28941282)
Metab Eng. 2017 Nov;44:1-12. (PMID: 28867349)
Appl Environ Microbiol. 2013 Jul;79(14):4192-8. (PMID: 23603671)
World J Microbiol Biotechnol. 2018 Sep 10;34(10):145. (PMID: 30203131)
Appl Environ Microbiol. 2010 Oct;76(19):6370-6. (PMID: 20709850)
Appl Microbiol Biotechnol. 2017 Jan;101(1):241-251. (PMID: 27734122)
World J Microbiol Biotechnol. 2018 Aug 20;34(9):135. (PMID: 30128628)
J Bacteriol. 2004 Mar;186(5):1475-83. (PMID: 14973018)
Appl Environ Microbiol. 2018 Oct 1;84(20):. (PMID: 30097440)
J Agric Food Chem. 2017 Mar 29;65(12):2540-2548. (PMID: 28262014)
J Appl Microbiol. 2016 Sep;121(3):704-12. (PMID: 27159567)
Microb Drug Resist. 1996 Spring;2(1):123-9. (PMID: 9158734)
ACS Synth Biol. 2019 Apr 19;8(4):866-875. (PMID: 30865822)

2018YFA0900303 National Key Research and Development Program of China; 2015CB150505 National Key Research and Development Program of China; 2018ACA149 Technical Innovation Special Fund of Hubei Province; 2018M642802 China Postdoctoral Science Foundation; 027Y2019-018 the Science and Technology Project of Hubei Tobacco Company

Keywords: Bacillus licheniformis; Cell surface engineering; Protein expression; clsA; dltD; pssA

0 (Membrane Proteins)
EC 2.7.8.- (Transferases (Other Substituted Phosphate Groups))
EC 2.7.8.- (cardiolipin synthetase)
EC 3.2.1.1 (alpha-Amylases)
EC 3.4.21.- (Subtilisins)
H81695M5OP (nattokinase)

Date Created: 20190901 Date Completed: 20200124 Latest Revision: 20200124

20221213

10.1007/s10295-019-02229-8

31471782