Abstract
We present a baculovirus-based protein engineering method that enables site-specific introduction of unique functionalities in a eukaryotic protein complex recombinantly produced in insect cells. We demonstrate the versatility of this efficient and robust protein production platform, 'MultiBacTAG', (i) for the fluorescent labeling of target proteins and biologics using click chemistries, (ii) for glycoengineering of antibodies, and (iii) for structure-function studies of novel eukaryotic complexes using single-molecule Förster resonance energy transfer as well as site-specific crosslinking strategies.
MeSH terms
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Animals
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Baculoviridae / genetics
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Baculoviridae / metabolism
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Cell Culture Techniques
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Fluorescence Resonance Energy Transfer / methods
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Genetic Code
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Genetic Vectors
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Green Fluorescent Proteins / biosynthesis*
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Green Fluorescent Proteins / chemistry
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Green Fluorescent Proteins / genetics
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Humans
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Multiprotein Complexes / biosynthesis*
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Multiprotein Complexes / chemistry
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Multiprotein Complexes / genetics
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Plasmids
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Protein Engineering / methods*
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Recombinant Proteins / biosynthesis*
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Recombinant Proteins / chemistry
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Recombinant Proteins / genetics
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Sf9 Cells
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Spodoptera
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Viral Proteins / biosynthesis*
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Viral Proteins / chemistry
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Viral Proteins / genetics
Substances
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Multiprotein Complexes
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Recombinant Proteins
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Viral Proteins
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Green Fluorescent Proteins