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Preparation of a genetically fused protein A/luciferase conjugate for use in bioluminescent immunoassays

Authors
Journal
Journal of Immunological Methods
0022-1759
Publisher
Elsevier
Publication Date
Volume
137
Issue
2
Identifiers
DOI: 10.1016/0022-1759(91)90025-b
Keywords
  • Protein A
  • Luciferase
  • Bioluminescence
  • Immunoassay
  • Fusion Protein
Disciplines
  • Biology

Abstract

Abstract The genes encoding staphylococcal protein A and bacterial luciferase ( Vibrio harveyi) were fused in-frame in order to obtain a general marker enzyme for bioluminescent immunoassays. Two constructs were made where protein A was ligated to the first and the 12th amino acid residue, respectively, of the N terminus of the β subunit of luciferase. Only the first fusion protein encoding the entire β subunit was able to form an enzymatically active luciferase complex when expressed together with the α subunit. The fusion of protein A to luciferase did not notably alter the emitted wavelength spectrum or its stability to urea treatment. The fusion protein was found to retain at least 50% of the specific bioluminescent activity compared to native luciferase. In preliminary tests, this hybrid protein was shown to be useful in bioluminescent immunoassays.

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