Two proteins for the price of one: structural studies of the dual-destiny protein preproalbumin with sunflower trypsin inhibitor-1

Franke, Bastian, James, Amy M., Mobli, Mehdi, Colgrave, Michelle L., Mylne, Joshua S. and Rosengren, K. Johan (2017) Two proteins for the price of one: structural studies of the dual-destiny protein preproalbumin with sunflower trypsin inhibitor-1. The Journal of Biological Chemistry, 292 30: 12398-12411. doi:10.1074/jbc.M117.776955

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Author Franke, Bastian
James, Amy M.
Mobli, Mehdi
Colgrave, Michelle L.
Mylne, Joshua S.
Rosengren, K. Johan
Title Two proteins for the price of one: structural studies of the dual-destiny protein preproalbumin with sunflower trypsin inhibitor-1
Journal name The Journal of Biological Chemistry   Check publisher's open access policy
ISSN 1083-351X
Publication date 2017-07-28
Year available 2017
Sub-type Article (original research)
DOI 10.1074/jbc.M117.776955
Open Access Status File (Publisher version)
Volume 292
Issue 30
Start page 12398
End page 12411
Total pages 14
Place of publication Rockville, MD United States
Publisher American Society for Biochemistry and Molecular Biology
Language eng
Subject 1303 Biochemistry
1312 Molecular Biology
1307 Cell Biology
Abstract Seed storage proteins are both an important source of nutrition for humans and essential for seedling establishment. Interestingly, unusual napin-type 2S seed storage albumin precursors in sunflowers contain a sequence that is released as a macrocyclic peptide during post-translational processing. The mechanism by which such peptides emerge from linear precursor proteins has received increased attention; however, the structural characterization of intact precursor proteins has been limited. Here, we report the 3D NMR structure of the Helianthus annuus PawS1 (preproalbumin with sunflower trypsin inhibitor-1) and provide new insights into the processing of this remarkable dual-destiny protein. In seeds, PawS1 is matured by asparaginyl endopeptidases (AEPs) into the cyclic peptide SFTI-1 (sunflower trypsin inhibitor-1) and a heterodimeric 2S albumin. The structure of PawS1 revealed that SFTI-1 and the albumin are independently folded into well-defined domains separated by a flexible linker. PawS1 was cleaved in vitro with recombinant sunflower HaAEP1 and in situ using a sunflower seed extract in a way that resembled the expected in vivo cleavages. Recombinant HaAEP1 cleaved PawS1 at multiple positions, and in situ, its flexible linker was removed, yielding fully mature heterodimeric albumin. Liberation and cyclization of SFTI-1, however, was inefficient, suggesting that specific seed conditions or components may be required for in vivo biosynthesis of SFTI-1. In summary, this study has revealed the 3D structure of a macrocyclic precursor protein and provided important mechanistic insights into the maturation of sunflower proalbumins into an albumin and a macrocyclic peptide.
Formatted abstract
Seed storage proteins are both an important source of nutrition for humans and essential for seedling establishment. Interestingly, unusual napin-type 2S seed storage albumin precursors in sunflowers contain a sequence that is released as a macrocyclic peptide during post-translational processing. The mechanism by which such peptides emerge from linear precursor proteins has received increased attention; however, the structural characterization of intact precursor proteins has been limited. Here, we report the 3D NMR structure of the Helianthus annuus PawS1 (preproalbumin with sunflower trypsin inhibitor-1) and provide new insights into the processing of this remarkable dual-destiny protein. In seeds, PawS1 is matured by asparaginyl endopeptidases (AEPs) into the cyclic peptide SFTI-1 (sunflower trypsin inhibitor-1) and a heterodimeric 2S albumin. The structure of PawS1 revealed that SFTI-1 and the albumin are independently folded into well-defined domains separated by a flexible linker. PawS1 was cleaved in vitro with recombinant sunflower HaAEP1 and in situ using a sunflower seed extract in a way that resembled the expected in vivo cleavages. Recombinant HaAEP1 cleaved PawS1 at multiple positions, and in situ, its flexible linker was removed, yielding fully mature heterodimeric albumin. Liberation and cyclization of SFTI-1, however, was inefficient, suggesting that specific seed conditions or components may be required for in vivo biosynthesis of SFTI-1. In summary, this study has revealed the 3D structure of a macrocyclic precursor protein and provided important mechanistic insights into the maturation of sunflower proalbumins into an albumin and a macrocyclic peptide.
Keyword asparaginyl endopeptidase (AEP)
cyclic peptide
nuclear magnetic resonance (NMR)
plant biochemistry
post-translational modification (PTM)
preproalbumin with SFTI-1 (PawS1)
protein processing
seed storage albumin
sunflower trypsin inhibitor-1 (SFTI-1)
Q-Index Code C1
Q-Index Status Provisional Code
Grant ID DP120103369
Institutional Status UQ

 
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