Strategies for enhancing bioluminescent bacterial sensor performance by promoter region manipulation.
Bioluminescent bacterial sensors are based upon the fusion of bacterial bioluminescence (lux) genes, acting as a reporter element, to selected bacterial stress-response gene promoters. Depending upon the nature of the promoter, the resulting constructs react to diverse types of environmental stress, including the presence of toxic chemicals, by dose-dependant light emission. Two bacterial sensors, harbouring sulA::luxCDABE and grpE::luxCDABE fusions, activated by the model chemicals nalidixic acid (NA) and ethanol, respectively, were subjected to molecular manipulations of the promoter region, in order to enhance the intensity and speed of their response and lower their detection thresholds. By manipulating the length of the promoter-containing segment (both promoters), by introducing random or specific mutations in the promoter sequence or by duplicating the promoter sequence (sulA only), major improvements in sensor performance were obtained. Improvements included significantly enhanced sensitivity, earlier response times and an increase in signal intensity. The general approaches described herein may be of general applicability for optimizing bacterial sensor performance, regardless of the sensing or reporting elements employed.
Institute of Life Sciences, The Hebrew University of Jerusalem, Jerusalem 91904, Israel.
Sharon
S
Yagur-Kroll
Benny
B
Bilic
Shimshon
S
Belkin
D001419:Bacteria / Q000235:genetics* / Q000378:metabolism*; D015374:Biosensing Techniques / Q000379:methods*; D008163:Luminescent Measurements / Q000379:methods*; D008164:Luminescent Proteins / Q000096:biosynthesis*; D009154:Mutation; D011401:Promoter Regions, Genetic
D008164:Luminescent Proteins
Environ Sci Technol. 2003 Oct 15;37(20):4743-50
14594387
Biotechnol Bioeng. 2007 Feb 15;96(3):550-8
16964624
Microb Biotechnol. 2008 Jul;1(4):320-30
21261850
J Biotechnol. 2005 Feb 9;115(3):239-48
15639086
Nucleic Acids Res. 1987 Mar 11;15(5):2343-61
3550697
J Bacteriol. 2001 May;183(9):2866-73
11292807
Appl Environ Microbiol. 1994 May;60(5):1414-20
8017928
Environ Microbiol. 2004 Nov;6(11):1186-96
15479251
J Mol Biol. 1994 Aug 26;241(4):507-23
8057377
Nucleic Acids Res. 1990 Dec 25;18(24):7367-72
2259628
J Biotechnol. 2006 Feb 10;121(3):291-8
16150509
J Mol Biol. 1983 Dec 15;171(3):337-43
6317868
Mutat Res. 2002 Jan 15;513(1-2):169-82
11719102
J Bacteriol. 2001 Sep;183(18):5441-4
11514531
Mutat Res. 1989 Oct;216(5):251-7
2677705
J Biosci Bioeng. 2001;92(3):301-4
16233101
Science. 1997 Sep 5;277(5331):1453-62
9278503
Nucleic Acids Res. 1993 Apr 11;21(7):1507-16
8479900
Appl Environ Microbiol. 2005 May;71(5):2338-46
15870320
Proc Natl Acad Sci U S A. 1982 Oct;79(19):5971-5
6821127
Nat Rev Microbiol. 2004 Jan;2(1):57-65
15035009
Methods Mol Biol. 1998;102:247-58
9680625
Proc Natl Acad Sci U S A. 1995 Apr 11;92(8):3526-30
7536935
Adv Biophys. 1986;21:163-73
3529848
Proc Natl Acad Sci U S A. 2005 Sep 6;102(36):12678-83
16123130
Microbiol Rev. 1984 Mar;48(1):60-93
6371470
Science. 2009 Apr 10;324(5924):255-8
19359587
Mutat Res. 1997 Mar 17;389(2-3):279-90
9093394
Science. 1990 Aug 17;249(4970):778-81
17756791
Microb Biotechnol. 2008 Mar;1(2):149-57
21261832
Protein Expr Purif. 2005 Jun;41(2):433-40
15866732
Nucleic Acids Res. 1983 Apr 25;11(8):2237-55
6344016
Biotechnol Prog. 1996 May-Jun;12(3):387-92
8652122
Protein Sci. 1993 Jul;2(7):1072-84
8102921
Nat Methods. 2006 Aug;3(8):623-8
16862137