title: Engineering foot-and-mouth disease virus serotype O IND R2/1975 for one-step purification by immobilized metal affinity chromatography.
abstract: Immobilized metal affinity chromatography (IMAC) allows for the efficient protein purification via metal affinity tag such as hexa-histidine (His6) sequence. To develop a new chromatography strategy for the purification and concentration of foot-and-mouth disease virus (FMDV) particles, we inserted the His6-tag at the earlier reported site in the VP1 G-H loop of the FMD virus serotype O vaccine strain IND R2/1975. Display of the His6-tag on the capsid surface, endowed the virus with an increased affinity for immobilized nickel ions. We demonstrated that the His6-tagged FMDV could be produced to high titre and purified from the infected BHK-21 cell lysates by IMAC efficiently. Further, a 1150-fold reduction in protein contaminant level and an 8400-fold reduction in DNA contaminant level were achieved in the IMAC purification of His6-tagged FMDV. Through various functional assays it has been found that the tagged virus retains its functionality and infectivity similar to the non-tagged virus. The affinity purification of the His6-tagged FMDV may offer a feasible, alternative approach to the current methods of FMDV antigen purification, concentration and process scalability.
authors.affiliation: ICAR-Project Directorate on Foot-and-Mouth Disease, Mukteswar, Nainital 263 138, Uttarakhand, India. Electronic address: jkubiswal@gmail.com.
authors.forename: Jitendra K
authors.initials: JK
authors.lastname: Biswal
authors.affiliation: ICAR-Project Directorate on Foot-and-Mouth Disease, Mukteswar, Nainital 263 138, Uttarakhand, India.
authors.forename: Punam
authors.initials: P
authors.lastname: Bisht
authors.affiliation: ICAR-Project Directorate on Foot-and-Mouth Disease, Mukteswar, Nainital 263 138, Uttarakhand, India.
authors.forename: Saravanan
authors.initials: S
authors.lastname: Subramaniam
authors.affiliation: ICAR-Project Directorate on Foot-and-Mouth Disease, Mukteswar, Nainital 263 138, Uttarakhand, India.
authors.forename: Rajeev
authors.initials: R
authors.lastname: Ranjan
authors.affiliation: ICAR-Project Directorate on Foot-and-Mouth Disease, Mukteswar, Nainital 263 138, Uttarakhand, India.
authors.forename: Gaurav K
authors.initials: GK
authors.lastname: Sharma
authors.affiliation: ICAR-Project Directorate on Foot-and-Mouth Disease, Mukteswar, Nainital 263 138, Uttarakhand, India. Electronic address: pattnaikb@gmail.com.
authors.forename: Bramhadev
authors.initials: B
authors.lastname: Pattnaik
mesh_terms: D000818:Animals; D002460:Cell Line; D002846:Chromatography, Affinity / Q000379:methods*; D006224:Cricetinae; D004279:DNA, Viral / Q000235:genetics; D029544:Foot-and-Mouth Disease Virus / Q000235:genetics / Q000302:isolation & purification* / Q000502:physiology; D009532:Nickel / Q000737:chemistry; D014779:Virus Replication
chemical_list: D004279:DNA, Viral; D009532:Nickel
keywords: Affinity chromatography; FMD virus; FMDV purification; His(6)-tagged FMDV; Reverse genetics