Title
Structural characterization of potato protease inhibitor I (cv. Bintje) after expression in Pichia pastoris
Author
TNO Zeist
van den Broek, L.A.M.
Pouvreau, L.
Lommerse, G.
Schipper, B.
van Koningsveld, G.A.
Gruppen, H.
Publication year
2004
Abstract
In the present study the structural properties of potato protease inhibitor 1 (PI-1) were studied as a function of temperature to elucidate its precipitation mechanism upon heating. A cDNA coding for PI-1 from cv. Bintje was cloned and expressed in Pichia pastoris. Using the recombinant PI-1 it was suggested that PI-1 behaves as a hexameric protein rather than as a pentamer, as previously proposed in the literature. The recombinant protein seems either to have a predominantly unordered structure or to belong to the β-II proteins. Differential scanning calorimetry analysis of PI-1 revealed that its thermal unfolding occurs via one endothermic transition in which the hexameric PI-1 probably unfolds, having a dimer instead of a monomer as cooperative unit. The transition temperature for the recombinant PI-1 was 88 °C. Similar results were obtained for a partially purified pool of native PI-1 from cv. Bintje. Molecular Sequence Numbers: GENBANK: AY496262; Chemicals / CAS: proteinase inhibitor, 37205-61-1; Plant Proteins; Protease Inhibitors; Recombinant Proteins
Subject
Potato
Protease inhibitor
Recombinant PI-1
Solanum tuberosum
complementary DNA
proteinase inhibitor 1
recombinant protein
unclassified drug
differential scanning calorimetry
enzyme inhibition
gene expression
nonhuman
nucleotide sequence
Pichia pastoris
polyacrylamide gel electrophoresis
polymerase chain reaction
protein purification
proteinase inhibition
root vegetable
Calorimetry, Differential Scanning
Chemistry, Physical
Circular Dichroism
Drug Stability
Gene Expression
Heat
Molecular Weight
Pichia
Plant Proteins
Protease Inhibitors
Protein Folding
Recombinant Proteins
Solanum tuberosum
Pichia
Pichia pastoris
Solanum
Solanum tuberosum
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DOI
https://doi.org/10.1021/jf049832x
TNO identifier
237906
ISSN
0021-8561
Source
Journal of Agricultural and Food Chemistry, 52 (52), 4928-4934
Document type
article