Prokaryotic expression and characterisation of recombinant M1 protein of an Indian H5N1 avian influenza virus

ZOHRA SYED; GOVINDARAJULU VENKATESH; HARSHAD V MURUGKAR; MEGHA KADAM BEDEKAR; SHANMUGASUNDARAM NAGARAJAN; SHIV CHANDRA DUBEY

doi:10.56093/ijans.v82i12.25653

Authors

ZOHRA SYED High Security Animal Disease Laboratory, Indian Veterinary Research Institute, Anand Nagar, Bhopal, Madhya Pradesh 462 021 India
GOVINDARAJULU VENKATESH High Security Animal Disease Laboratory, Indian Veterinary Research Institute, Anand Nagar, Bhopal, Madhya Pradesh 462 021 India
HARSHAD V MURUGKAR High Security Animal Disease Laboratory, Indian Veterinary Research Institute, Anand Nagar, Bhopal, Madhya Pradesh 462 021 India
MEGHA KADAM BEDEKAR High Security Animal Disease Laboratory, Indian Veterinary Research Institute, Anand Nagar, Bhopal, Madhya Pradesh 462 021 India
SHANMUGASUNDARAM NAGARAJAN High Security Animal Disease Laboratory, Indian Veterinary Research Institute, Anand Nagar, Bhopal, Madhya Pradesh 462 021 India
SHIV CHANDRA DUBEY High Security Animal Disease Laboratory, Indian Veterinary Research Institute, Anand Nagar, Bhopal, Madhya Pradesh 462 021 India

https://doi.org/10.56093/ijans.v82i12.25653

Keywords:

Avian influenza, Characterization, Expression, Purification, Recombinant M1 protein

Abstract

The type specific Matrix 1 (M1) protein of an Indian H5N1 avian influenza virus (AIV) was expressed as a histidine- tagged fusion protein in a prokaryotic expression system and characterized. The M1 gene was amplified by reverse transcription PCR using appropriately designed primers and cloned into the expression vector, pET28a (+). The orientation and reading frame of the recombinant expression construct (pET-M1) was confirmed by sequence analysis. The derived amino acid sequence homology between M1 of AIV H5N1 and other reference AIV subtypes was found to be 93.7% to 99.2%. The 33kDA recombinant M1 (rM1) protein was expressed as inclusion body after induction with 1 mM IPTG in E. coli BL₂₁ (DE3)pLysS cells. The protein was purified to near homogeneity by affinity chromatography using Ni- NTA agarose column. The yield of the purified rM1 was found to be 2 mg/100 ml of induced culture. The rM1 was found to react specifically with H1-H15 AIV subtype specific sera in Western blot. The results indicated that the purified rM1 protein could be used as antigen for detection of type specific AIV antibodies by immunoassays.

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Author Biographies

ZOHRA SYED, High Security Animal Disease Laboratory, Indian Veterinary Research Institute, Anand Nagar, Bhopal, Madhya Pradesh 462 021 India
GOVINDARAJULU VENKATESH, High Security Animal Disease Laboratory, Indian Veterinary Research Institute, Anand Nagar, Bhopal, Madhya Pradesh 462 021 India
HARSHAD V MURUGKAR, High Security Animal Disease Laboratory, Indian Veterinary Research Institute, Anand Nagar, Bhopal, Madhya Pradesh 462 021 India
MEGHA KADAM BEDEKAR, High Security Animal Disease Laboratory, Indian Veterinary Research Institute, Anand Nagar, Bhopal, Madhya Pradesh 462 021 India
SHANMUGASUNDARAM NAGARAJAN, High Security Animal Disease Laboratory, Indian Veterinary Research Institute, Anand Nagar, Bhopal, Madhya Pradesh 462 021 India
SHIV CHANDRA DUBEY, High Security Animal Disease Laboratory, Indian Veterinary Research Institute, Anand Nagar, Bhopal, Madhya Pradesh 462 021 India

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