Structural and immunological characterization of E. coli derived recombinant CRM197 protein used as carrier in conjugate vaccines

Ravi P.N. Mishra, Ravi S.P. Yadav, Christopher Jones, Salvatore Nocadello, George Minasov, Ludmilla A Shuvalova, Wayne F Anderson, Akshay Goel*

*Corresponding author for this work

Research output: Contribution to journalArticle

Abstract

It is established that the immunogenicity of polysaccharides is enhanced by coupling them to carrier proteins. Cross reacting material (CRM197), a nontoxic variant of diphtheria toxin (DT) is widely used carrier protein for polysaccharide conjugate vaccines. Conventionally, CRM197 is isolated by fermentation of Corynebacterium diphtheriae C7 (β197) cultures, which often suffers from low yield. Recently, several recombinant approaches have been reported with robust processes and higher yields, which will improve the affordability of CRM197-based vaccines. Vaccine manufacturers require detailed analytical information to ensure that the CRM197 meets quality standards and regulatory requirements. In the present manuscript we have described detailed structural characteristics of Escherichia coli based recombinant CRM197 (rCRM197) carrier protein. The crystal structure of the E. coli based rCRM197 was found to be identical with the reported crystal structure of the C7 CRM197 produced in C. diphtheriae C7 strain (Protein Data Bank (PDB) ID: 4EA0). The crystal structure of rCRM197 was determined at 2.3 Å resolution and structure was submitted to the PDB with accession number ID 5I82. This is the first report of a crystal structure of E. coli derived recombinant CRM197 carrier protein. Furthermore, the rCRM197 was conjugated to Vi polysaccharide to generate Typhoid conjugate vaccine (Vi-rCRM197) and its immunogenicity was evaluated in Balb/C Mice. The Vi-rCRM197 conjugate vaccine was found to generate strong primary α-Vi antibody response and also showed a booster response after subsequent vaccination in mice. Overall data suggest that E. coli based recombinant CRM197 exhibits structural and immunological similarity with the C7 CRM197 and can be used as a carrier protein in conjugate vaccine development.

Original languageEnglish (US)
Article numberBSR20180238
JournalBioscience Reports
Volume38
Issue number5
DOIs
StatePublished - Sep 25 2018

Fingerprint

Recombinant proteins
Conjugate Vaccines
Recombinant Proteins
Escherichia coli
Carrier Proteins
Crystal structure
Corynebacterium diphtheriae
Polysaccharides
CRM197 (non-toxic variant of diphtheria toxin)
Vaccines
Typhoid-Paratyphoid Vaccines
Databases
Diphtheria Toxin
Manuscripts
Fermentation
Antibody Formation

ASJC Scopus subject areas

  • Biophysics
  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

@article{9a5961a4066045c2a15b0ffffaf6c7e8,
title = "Structural and immunological characterization of E. coli derived recombinant CRM197 protein used as carrier in conjugate vaccines",
abstract = "It is established that the immunogenicity of polysaccharides is enhanced by coupling them to carrier proteins. Cross reacting material (CRM197), a nontoxic variant of diphtheria toxin (DT) is widely used carrier protein for polysaccharide conjugate vaccines. Conventionally, CRM197 is isolated by fermentation of Corynebacterium diphtheriae C7 (β197) cultures, which often suffers from low yield. Recently, several recombinant approaches have been reported with robust processes and higher yields, which will improve the affordability of CRM197-based vaccines. Vaccine manufacturers require detailed analytical information to ensure that the CRM197 meets quality standards and regulatory requirements. In the present manuscript we have described detailed structural characteristics of Escherichia coli based recombinant CRM197 (rCRM197) carrier protein. The crystal structure of the E. coli based rCRM197 was found to be identical with the reported crystal structure of the C7 CRM197 produced in C. diphtheriae C7 strain (Protein Data Bank (PDB) ID: 4EA0). The crystal structure of rCRM197 was determined at 2.3 {\AA} resolution and structure was submitted to the PDB with accession number ID 5I82. This is the first report of a crystal structure of E. coli derived recombinant CRM197 carrier protein. Furthermore, the rCRM197 was conjugated to Vi polysaccharide to generate Typhoid conjugate vaccine (Vi-rCRM197) and its immunogenicity was evaluated in Balb/C Mice. The Vi-rCRM197 conjugate vaccine was found to generate strong primary α-Vi antibody response and also showed a booster response after subsequent vaccination in mice. Overall data suggest that E. coli based recombinant CRM197 exhibits structural and immunological similarity with the C7 CRM197 and can be used as a carrier protein in conjugate vaccine development.",
author = "Mishra, {Ravi P.N.} and Yadav, {Ravi S.P.} and Christopher Jones and Salvatore Nocadello and George Minasov and Shuvalova, {Ludmilla A} and Anderson, {Wayne F} and Akshay Goel",
year = "2018",
month = "9",
day = "25",
doi = "10.1042/BSR20180238",
language = "English (US)",
volume = "38",
journal = "Bioscience Reports",
issn = "0144-8463",
publisher = "Portland Press Ltd.",
number = "5",

}

Structural and immunological characterization of E. coli derived recombinant CRM197 protein used as carrier in conjugate vaccines. / Mishra, Ravi P.N.; Yadav, Ravi S.P.; Jones, Christopher; Nocadello, Salvatore; Minasov, George; Shuvalova, Ludmilla A; Anderson, Wayne F; Goel, Akshay.

In: Bioscience Reports, Vol. 38, No. 5, BSR20180238, 25.09.2018.

Research output: Contribution to journalArticle

TY - JOUR

T1 - Structural and immunological characterization of E. coli derived recombinant CRM197 protein used as carrier in conjugate vaccines

AU - Mishra, Ravi P.N.

AU - Yadav, Ravi S.P.

AU - Jones, Christopher

AU - Nocadello, Salvatore

AU - Minasov, George

AU - Shuvalova, Ludmilla A

AU - Anderson, Wayne F

AU - Goel, Akshay

PY - 2018/9/25

Y1 - 2018/9/25

N2 - It is established that the immunogenicity of polysaccharides is enhanced by coupling them to carrier proteins. Cross reacting material (CRM197), a nontoxic variant of diphtheria toxin (DT) is widely used carrier protein for polysaccharide conjugate vaccines. Conventionally, CRM197 is isolated by fermentation of Corynebacterium diphtheriae C7 (β197) cultures, which often suffers from low yield. Recently, several recombinant approaches have been reported with robust processes and higher yields, which will improve the affordability of CRM197-based vaccines. Vaccine manufacturers require detailed analytical information to ensure that the CRM197 meets quality standards and regulatory requirements. In the present manuscript we have described detailed structural characteristics of Escherichia coli based recombinant CRM197 (rCRM197) carrier protein. The crystal structure of the E. coli based rCRM197 was found to be identical with the reported crystal structure of the C7 CRM197 produced in C. diphtheriae C7 strain (Protein Data Bank (PDB) ID: 4EA0). The crystal structure of rCRM197 was determined at 2.3 Å resolution and structure was submitted to the PDB with accession number ID 5I82. This is the first report of a crystal structure of E. coli derived recombinant CRM197 carrier protein. Furthermore, the rCRM197 was conjugated to Vi polysaccharide to generate Typhoid conjugate vaccine (Vi-rCRM197) and its immunogenicity was evaluated in Balb/C Mice. The Vi-rCRM197 conjugate vaccine was found to generate strong primary α-Vi antibody response and also showed a booster response after subsequent vaccination in mice. Overall data suggest that E. coli based recombinant CRM197 exhibits structural and immunological similarity with the C7 CRM197 and can be used as a carrier protein in conjugate vaccine development.

AB - It is established that the immunogenicity of polysaccharides is enhanced by coupling them to carrier proteins. Cross reacting material (CRM197), a nontoxic variant of diphtheria toxin (DT) is widely used carrier protein for polysaccharide conjugate vaccines. Conventionally, CRM197 is isolated by fermentation of Corynebacterium diphtheriae C7 (β197) cultures, which often suffers from low yield. Recently, several recombinant approaches have been reported with robust processes and higher yields, which will improve the affordability of CRM197-based vaccines. Vaccine manufacturers require detailed analytical information to ensure that the CRM197 meets quality standards and regulatory requirements. In the present manuscript we have described detailed structural characteristics of Escherichia coli based recombinant CRM197 (rCRM197) carrier protein. The crystal structure of the E. coli based rCRM197 was found to be identical with the reported crystal structure of the C7 CRM197 produced in C. diphtheriae C7 strain (Protein Data Bank (PDB) ID: 4EA0). The crystal structure of rCRM197 was determined at 2.3 Å resolution and structure was submitted to the PDB with accession number ID 5I82. This is the first report of a crystal structure of E. coli derived recombinant CRM197 carrier protein. Furthermore, the rCRM197 was conjugated to Vi polysaccharide to generate Typhoid conjugate vaccine (Vi-rCRM197) and its immunogenicity was evaluated in Balb/C Mice. The Vi-rCRM197 conjugate vaccine was found to generate strong primary α-Vi antibody response and also showed a booster response after subsequent vaccination in mice. Overall data suggest that E. coli based recombinant CRM197 exhibits structural and immunological similarity with the C7 CRM197 and can be used as a carrier protein in conjugate vaccine development.

UR - http://www.scopus.com/inward/record.url?scp=85054240303&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=85054240303&partnerID=8YFLogxK

U2 - 10.1042/BSR20180238

DO - 10.1042/BSR20180238

M3 - Article

C2 - 29875175

AN - SCOPUS:85054240303

VL - 38

JO - Bioscience Reports

JF - Bioscience Reports

SN - 0144-8463

IS - 5

M1 - BSR20180238

ER -