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Analysis of an autoproteolytic activity of rice yellow mottle virus silencing suppressor P1

Ectopically expressed rice yellow mottle virus P1 fusion proteins were found to be cleaved in planta and in Escherichia coli . Cleavage takes place in the absence of bacterial protease activity, indicating that the P1 fusion is autocatalytically processed independently of host factors. N-terminal se...

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Main Authors: Weinheimer, Isabel (Author) , Kajohn Boonrod (Author) , Moser, Mirko (Author) , Zwiebel, Michèle (Author) , Füllgrabe, Marc W. (Author) , Krczal, Gabi (Author) , Wassenegger, Michael (Author)
Format: Article (Journal)
Language:English
Published: 26. Februar 2010
In: Biological chemistry
Year: 2010, Volume: 391, Issue: 2-3, Pages: 271-281
ISSN:1437-4315
DOI:10.1515/bc.2010.022
Online Access:Resolving-System, lizenzpflichtig, Volltext: https://doi.org/10.1515/bc.2010.022
Verlag, lizenzpflichtig, Volltext: https://www.degruyterbrill.com/document/doi/10.1515/bc.2010.022/html
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Author Notes:Isabel Weinheimer, Kajohn Boonrod, Mirko Moser, Michèle Zwiebel, Marc Füllgrabe, Gabi Krczal and Michael Wassenegger
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Summary:Ectopically expressed rice yellow mottle virus P1 fusion proteins were found to be cleaved in planta and in Escherichia coli . Cleavage takes place in the absence of bacterial protease activity, indicating that the P1 fusion is autocatalytically processed independently of host factors. N-terminal sequencing of the C-terminal cleavage product of transiently expressed P1/GFP (green fluorescence protein) in Nicotiana benthamiana showed that the cleavage site is located between the first two amino acids (aa) downstream of the P1 sequence. Mutagenesis experiments revealed that a phenylalanine to valine substitution at position 157 of the P1 aa sequence impairs proper cleavage, which is nearly unaffected by replacement of phenylalanine with tyrosine. Deletion of methionine 159 (first GFP aa residue) appeared to not affect P1/GFP cleavage. N-terminal P1-tagging with GFP turned out to impair autocleavage, whereas a small His-tag could not fully prevent cleavage. Additionally, a modified P1/GFP carrying an N-terminal deletion of 81 aa was not cleaved. These findings indicate that this region is involved in the proteolysis mechanism and that large N-terminal fusion partners might affect correct folding of the P1 necessary for self-catalysis.
Item Description:Gesehen am 19.10.2023
Physical Description:Online Resource
ISSN:1437-4315
DOI:10.1515/bc.2010.022

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