Structural modifications of outer membrane vesicles to refine them as vaccine delivery vehicles.
In an effort to devise a safer and more effective vaccine delivery system, outer membrane vesicles (OMVs) were engineered to have properties of intrinsically low endotoxicity sufficient for the delivery of foreign antigens. Our strategy involved mutational inactivation of the MsbB (LpxM) lipid A acyltransferase to generate OMVs of reduced endotoxicity from Escherichia coli (E. coli) O157:H7. The chromosomal tagging of a foreign FLAG epitope within an OmpA-fused protein was exploited to localize the FLAG epitope in the OMVs produced by the E. coli mutant having the defined msbB and the ompA::FLAG mutations. It was confirmed that the desired fusion protein (OmpA::FLAG) was expressed and destined to the outer membrane (OM) of the E. coli mutant from which the OMVs carrying OmpA::FLAG are released during growth. A luminal localization of the FLAG epitope within the OMVs was inferred from its differential immunoprecipitation and resistance to proteolytic degradation. Thus, by using genetic engineering-based approaches, the native OMVs were modified to have both intrinsically low endotoxicity and a foreign epitope tag to establish a platform technology for development of multifunctional vaccine delivery vehicles.
Kim
Sang-Hyun
SH
Korea Research Institute of Bioscience and Biotechnology, Ochang, Cheongwon, Chungbuk 363-883, Republic of Korea.
Kim
Keun-Su
KS
Lee
Sang-Rae
SR
Kim
Ekyune
E
Kim
Myeong-Su
MS
Lee
Eun-Young
EY
Gho
Yong Song
YS
Kim
Jung-Woo
JW
Bishop
Russell E
RE
Chang
Kyu-Tae
KT
D000217:Acyltransferases / Q000235:genetics* / Q000378:metabolism*; D000595:Amino Acid Sequence; D000942:Antigens, Bacterial / Q000378:metabolism; D002855:Chromatography, Thin Layer; D000939:Epitopes; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000235:genetics* / Q000378:metabolism*; D005818:Genetic Engineering; D047468:Immunoprecipitation; D008050:Lipid A / Q000378:metabolism; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D022142:Secretory Vesicles / Q000737:chemistry* / Q000378:metabolism
D000942:Antigens, Bacterial; D000939:Epitopes; D029968:Escherichia coli Proteins; D008050:Lipid A; D011993:Recombinant Fusion Proteins; D000217:Acyltransferases; C097895:MsbB protein, E coli
Int Immunol. 2005 Jun;17(6):695-704
15843436
Infect Immun. 2004 Feb;72(2):1174-80
14742570
Nat Immunol. 2002 Apr;3(4):354-9
11912497
J Mol Biol. 2007 Oct 26;373(3):529-40
17868697
Infect Immun. 2008 Oct;76(10):4554-63
18678672
Biochim Biophys Acta. 1999 Sep 21;1421(1):77-90
10561473
Infect Immun. 1999 Dec;67(12):6583-90
10569778
Proteomics. 2007 Sep;7(17):3143-53
17787032
J Biol Chem. 1999 Jun 25;274(26):18503-14
10373459
J Biol Chem. 2005 Nov 18;280(46):38383-94
16103114
Microbiology (Reading). 2006 Mar;152(Pt 3):657-666
16514146
Infect Immun. 2001 Oct;69(10):5981-90
11553534
Nat Struct Biol. 1998 Nov;5(11):1013-7
9808047
Mol Microbiol. 2005 Aug;57(4):900-12
16091033
J Biol Chem. 2008 Sep 26;283(39):26748-58
18641391
J Immunol. 2000 Nov 1;165(9):5177-85
11046050
J Clin Invest. 1996 Jan 15;97(2):359-65
8567955
J Biol Chem. 2008 Feb 15;283(7):4332-43
18070877
DNA Res. 2001 Feb 28;8(1):11-22
11258796
J Biol Chem. 1997 Apr 18;272(16):10353-60
9099672
Infect Immun. 2003 Nov;71(11):6132-40
14573628
Vaccine. 2005 May 31;23(29):3762-74
15893613
Proc Natl Acad Sci U S A. 2000 Jun 6;97(12):6640-5
10829079
J Immunol. 2007 Dec 1;179(11):7692-701
18025215
Microbiol Mol Biol Rev. 2003 Dec;67(4):593-656
14665678
Mol Gen Genet. 1994 Apr;243(2):127-35
8177210
J Bacteriol. 1999 Aug;181(16):4725-33
10438737
Annu Rev Biochem. 2002;71:635-700
12045108
Immunity. 2005 May;22(5):551-60
15894273
Infect Immun. 2003 Sep;71(9):4985-95
12933841
Microbiology (Reading). 2007 Nov;153(Pt 11):3776-3784
17975086
Eur J Biochem. 1982 Feb;122(1):223-31
7037401
Methods. 1999 Sep;19(1):174-86
10525454
Annu Rev Biochem. 2007;76:295-329
17362200
Infect Immun. 2006 May;74(5):2676-85
16622204
Genes Dev. 2005 Nov 15;19(22):2645-55
16291643
J Bacteriol. 1983 Apr;154(1):269-77
6187729