Expression of the major bean proteins from Theobroma cacao (cocoa) in the yeasts Hansenula polymorpha and Saccharomyces cerevisiae
Küçük Resim Yok
Tarih
1996
Dergi Başlığı
Dergi ISSN
Cilt Başlığı
Yayıncı
Elsevier Science B.V., Amsterdam
Erişim Hakkı
info:eu-repo/semantics/closedAccess
Özet
The production in two yeast expression systems of recombinant forms of the major proteins from the cocoa bean is described. Three major protein species are found in the cocoa bean: an albumin of molecular mass 21 kDa (p21) and two insoluble vicilin-like proteins of molecular mass 31 kDa and 47 kDa (p31 and p47, respectively). The p31 and p47 species are known to be derived from a common 67-kDa precursor (p67) by post-translational processing that includes the deletion of a hydrophilic domain located immediately after an N-terminal signal sequence. All three proteins appear to be targeted to membrane-bound storage organelles by N-terminal signal sequences. The p21 and p67 coding sequences were expressed in Hansenula polymorpha using the powerful methanol oxidase (MOX) promoter and in Saccharomyces cerevisiae using the promoter of the pyruvate kinase (PYK) gene. The expression constructs contained the native plant signal sequence, or various yeast signals. The p21 protein was successfully expressed and secreted from both yeasts. The insoluble p67 protein proved more difficult. Species of the correct molecular mass were recovered internally and small amounts of a p47 species were secreted using a yeast leader sequence. However, proteolytic cleavage, probably due to Kex2p-like processing, led to the appearance of other protein species.The production in two yeast expression systems of recombinant forms of the major proteins from the cocoa bean is described. Three major protein species are found in the cocoa bean: an albumin of molecular mass 21 kDa (p21) and two insoluble vicilin-like proteins of molecular mass 31 kDa and 47 kDa (p31 and p47, respectively). The p31 and p47 species me known to be derived from a common 67-kDa precursor (p67) by post-translational processing that includes the deletion of a hydrophilic domain located immediately after an N-terminal signal sequence. All three proteins appear to be targeted to membrane-bound storage organelles by N-terminal signal sequences. The p21 and p67 coding sequences were expressed in Hansenula polymorpha using the powerful methanol oxidase (MOX) promoter and in Saccharomyces cerevisiae using the promoter of the pyruvate kinase (PYK) gene. The expression constructs contained the native plant signal sequence, or various yeast signals. The p21 protein was successfully expressed and secreted from both yeasts. The insoluble p67 protein proved more difficult. Species of the correct molecular mass were recovered internally and small amounts of a p47 species were secreted using a yeast leader sequence. However, proteolytic cleavage, probably due to Kex2p-like processing, led to the appearance of other protein species.
Açıklama
Anahtar Kelimeler
Hansenula Polymorpha; Mox Promoter; Pyk Promoter; Recombinant Cocoa Protein; Saccharomyces Cerevisiae; Theobroma Cacao, Biotechnology; Enzymes; Genes; Plants (Botany); Yeast; Albumin; Bean Proteins; Hansenula Polymorpha; Saccharomyces Cerevisiae; Theobroma Cacao; Viciline Like Protein; Yeast Expression; Proteins; Recombinant Protein; Article; Cacao; Controlled Study; Gene Expression Regulation; Hansenula Polymorpha; Molecular Cloning; Nonhuman; Priority Journal; Promoter Region; Saccharomyces Cerevisiae; Yeast; Phaseolus (Angiosperm); Pichia Angusta; Saccharomyces Cerevisiae; Theobroma Cacao; Theobroma Cacao
Kaynak
Journal of Biotechnology
WoS Q Değeri
Scopus Q Değeri
Q2
Cilt
46
Sayı
1
