Efficient production and characterization of the sweet-tasting brazzein secreted by the yeast [i]Pichia pastoris[/i] - Archive ouverte HAL Accéder directement au contenu
Article Dans Une Revue Journal of Agricultural and Food Chemistry Année : 2012

Efficient production and characterization of the sweet-tasting brazzein secreted by the yeast [i]Pichia pastoris[/i]

Résumé

Brazzein is a small, heat-, and pH-stable sweet protein present in the fruits of the West African plant Pentadiplandra brazzeana Baillon. It exists in two forms differing in sweetness intensity. The major form, called pyrE-bra, contains a pyroglutamic acid at its N-terminus, while the minor form, called des-pyrE-bra, lacks this residue. Here we describe the heterologous expression in the methylotrophic yeast Pichia pastoris of two natural forms of brazzein, pyrE-bra and des-pyrE-bra, and an additional form, called Q1-bra, which is not naturally occurring in the fruit. Q1-bra differs from pyrE-bra in having a glutamine residue instead of pyrE at its N-terminus. Over an expression period of 6 days, we obtained approximately 90, 30, and 90 mg/L of purified recombinant pyrE-bra, Q1-bra, and des-pyrE-bra brazzein forms, respectively. Recombinant proteins were purified and submitted to mass spectrometry. and H-1 NMR spectroscopy. The data indicate that the recombinant brazzein forms were properly folded. Moreover, they activated the human sweet receptor in vitro and evoked sweetness in vivo with properties similar to those of the two natural brazzein forms.
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Dates et versions

hal-01223562 , version 1 (02-11-2015)

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Nicolas Poirier, Natacha Roudnitzky, Anne Brockhoff, Christine Belloir, Marie Maison, et al.. Efficient production and characterization of the sweet-tasting brazzein secreted by the yeast [i]Pichia pastoris[/i]. Journal of Agricultural and Food Chemistry, 2012, 60 (39), pp.9807 - 9814. ⟨10.1021/jf301600m⟩. ⟨hal-01223562⟩
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