Micoli F, Rondini S, Gavini M, Pisoni I, Lanzilao L, Colucci AM, Giannelli C, Pippi F, Sollai L, Pinto V, Berti F, Maclennan CA, Martin LB, Saul A A scalable method for O-antigen purification applied to different Salmonella serovars Analytical Biochemistry434(1) (2012)
136-145
Hepp-(1-7)-+
|
Subst-(1-4)-+ a-D-Galp-(1-6)-+ | EtN-(1---P---P---4)-+
| | | |
a-D-GlcpNAc-(1-2)-a-D-Glcp-(1-2)-a-D-Galp-(1-3)-D-Glcp-(1-3)-L-gro-a-D-manHepp-(1-3)-L-gro-a-D-manHepp-(1-5)-Kdo
|
P-?)-+
Subst = O-antigen (ID 28461, ID 28462, ID 28463)
NCBI PubMed ID:23142430 Publication DOI:10.1016/j.ab.2012.10.038 Journal NLM ID:0370535 Publisher: Academic Press Correspondence: francesca.micolinovartis.com Institutions: Novartis Vaccines Institute for Global Health, 53100 Siena, Italy, Novartis Vaccines and Diagnostics, 53100 Siena, Italy, MRC Centre for Immune Regulation, School of Immunity and Infection, College of Medicine and Dental Sciences, University of Birmingham, Birmingham B15 2TT, UK
The surface lipopolysaccharide of Gram-negative bacteria is both a virulence factor and a B-cell antigen. Antibodies against O-antigen of lipopolysaccharide may confer protection against infection, and O-antigen conjugates have been designed against multiple pathogens. Here, we describe simplified methodology for extraction and purification of O-antigen-core portion of Salmonella lipopolysaccharide, suitable for large scale production. Lipopolysaccharide extraction and delipidation is performed by acetic acid hydrolysis of whole bacterial culture, and can take place directly in a bioreactor, without previous isolation and inactivation of bacteria. Further O-antigen-core purification consists of rapid filtration and precipitation steps, without using enzymes or hazardous chemicals. The process was successfully applied to different Salmonella enterica serovars (Paratyphi A, Typhimurium and Enteritidis), obtaining good yields of high quality material, suitable for conjugate vaccines preparations.
Micoli F, Rondini S, Gavini M, Pisoni I, Lanzilao L, Colucci AM, Giannelli C, Pippi F, Sollai L, Pinto V, Berti F, Maclennan CA, Martin LB, Saul A A scalable method for O-antigen purification applied to different Salmonella serovars Analytical Biochemistry434(1) (2012)
136-145
NCBI PubMed ID:23142430 Publication DOI:10.1016/j.ab.2012.10.038 Journal NLM ID:0370535 Publisher: Academic Press Correspondence: francesca.micolinovartis.com Institutions: Novartis Vaccines Institute for Global Health, 53100 Siena, Italy, Novartis Vaccines and Diagnostics, 53100 Siena, Italy, MRC Centre for Immune Regulation, School of Immunity and Infection, College of Medicine and Dental Sciences, University of Birmingham, Birmingham B15 2TT, UK
The surface lipopolysaccharide of Gram-negative bacteria is both a virulence factor and a B-cell antigen. Antibodies against O-antigen of lipopolysaccharide may confer protection against infection, and O-antigen conjugates have been designed against multiple pathogens. Here, we describe simplified methodology for extraction and purification of O-antigen-core portion of Salmonella lipopolysaccharide, suitable for large scale production. Lipopolysaccharide extraction and delipidation is performed by acetic acid hydrolysis of whole bacterial culture, and can take place directly in a bioreactor, without previous isolation and inactivation of bacteria. Further O-antigen-core purification consists of rapid filtration and precipitation steps, without using enzymes or hazardous chemicals. The process was successfully applied to different Salmonella enterica serovars (Paratyphi A, Typhimurium and Enteritidis), obtaining good yields of high quality material, suitable for conjugate vaccines preparations.
Micoli F, Rondini S, Gavini M, Pisoni I, Lanzilao L, Colucci AM, Giannelli C, Pippi F, Sollai L, Pinto V, Berti F, Maclennan CA, Martin LB, Saul A A scalable method for O-antigen purification applied to different Salmonella serovars Analytical Biochemistry434(1) (2012)
136-145
NCBI PubMed ID:23142430 Publication DOI:10.1016/j.ab.2012.10.038 Journal NLM ID:0370535 Publisher: Academic Press Correspondence: francesca.micolinovartis.com Institutions: Novartis Vaccines Institute for Global Health, 53100 Siena, Italy, Novartis Vaccines and Diagnostics, 53100 Siena, Italy, MRC Centre for Immune Regulation, School of Immunity and Infection, College of Medicine and Dental Sciences, University of Birmingham, Birmingham B15 2TT, UK
The surface lipopolysaccharide of Gram-negative bacteria is both a virulence factor and a B-cell antigen. Antibodies against O-antigen of lipopolysaccharide may confer protection against infection, and O-antigen conjugates have been designed against multiple pathogens. Here, we describe simplified methodology for extraction and purification of O-antigen-core portion of Salmonella lipopolysaccharide, suitable for large scale production. Lipopolysaccharide extraction and delipidation is performed by acetic acid hydrolysis of whole bacterial culture, and can take place directly in a bioreactor, without previous isolation and inactivation of bacteria. Further O-antigen-core purification consists of rapid filtration and precipitation steps, without using enzymes or hazardous chemicals. The process was successfully applied to different Salmonella enterica serovars (Paratyphi A, Typhimurium and Enteritidis), obtaining good yields of high quality material, suitable for conjugate vaccines preparations.
Methods: 1H NMR, sugar analysis, HPLC, HPSEC, HPAEC-PAD, extraction, HR-MAS NMR, delipidation, acid hydrolyssis Comments, role: secondary modifications ([aDGlcp]n at C4 aDGalp residues) may be present according to strain and/or phage lysogeny; an O-antigen region to ID 27252
Related record ID(s): 27252, 28461, 28463 NCBI Taxonomy refs (TaxIDs):149539 Reference(s) to other database(s): GTC:G19543LK Show glycosyltransferases
Micoli F, Rondini S, Gavini M, Pisoni I, Lanzilao L, Colucci AM, Giannelli C, Pippi F, Sollai L, Pinto V, Berti F, Maclennan CA, Martin LB, Saul A A scalable method for O-antigen purification applied to different Salmonella serovars Analytical Biochemistry434(1) (2012)
136-145
a-Abep2(%)Ac-(1-3)-+ /Variants 0/-+
| |
-2)-a-D-Manp-(1-4)-a-L-Rhap-(1-3)-a-D-Galp-(1-
/Variants 0/ is:
?%a-D-Glcp-(1-6)-
OR (exclusively)
?%a-D-Glcp-(1-4)-
NCBI PubMed ID:23142430 Publication DOI:10.1016/j.ab.2012.10.038 Journal NLM ID:0370535 Publisher: Academic Press Correspondence: francesca.micolinovartis.com Institutions: Novartis Vaccines Institute for Global Health, 53100 Siena, Italy, Novartis Vaccines and Diagnostics, 53100 Siena, Italy, MRC Centre for Immune Regulation, School of Immunity and Infection, College of Medicine and Dental Sciences, University of Birmingham, Birmingham B15 2TT, UK
The surface lipopolysaccharide of Gram-negative bacteria is both a virulence factor and a B-cell antigen. Antibodies against O-antigen of lipopolysaccharide may confer protection against infection, and O-antigen conjugates have been designed against multiple pathogens. Here, we describe simplified methodology for extraction and purification of O-antigen-core portion of Salmonella lipopolysaccharide, suitable for large scale production. Lipopolysaccharide extraction and delipidation is performed by acetic acid hydrolysis of whole bacterial culture, and can take place directly in a bioreactor, without previous isolation and inactivation of bacteria. Further O-antigen-core purification consists of rapid filtration and precipitation steps, without using enzymes or hazardous chemicals. The process was successfully applied to different Salmonella enterica serovars (Paratyphi A, Typhimurium and Enteritidis), obtaining good yields of high quality material, suitable for conjugate vaccines preparations.
Methods: 1H NMR, sugar analysis, HPLC, HPSEC, HPAEC-PAD, extraction, HR-MAS NMR, delipidation, acid hydrolyssis Comments, role: Salmonella enterica serovar Typhimurium serogroup O:4,5. Secondary modifications (OAc at C2 aDAbe and aDGlcp at C4 or at C6 aDGalp residues) that may or may not be present according to strain and/or phage lysogeny; an O-antigen region to ID 27252
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