Extracellular production of riboflavin-binding protein, a potential bitter inhibitor, by Brevibacillus choshinensis

Riboflavin-binding protein (RBP) is a glycophosphoprotein found in hen eggs. We previously identified the extraordinary characteristic of RBP in reducing bitterness. For a more detailed study on the mode of action and industrial application of this characteristic, we investigated the microbial produ...

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Bibliographic Details
Published in:Protein expression and purification 2010-05, Vol.71 (1), p.85-90
Main Authors: Maehashi, Kenji, Matano, Mami, Saito, Makiko, Udaka, Shigezo
Format: Article
Language:English
Subjects:
Animals
Biochemistry - methods
Bitterness reduction
Brevibacillus choshinensis
Brevibacterium - metabolism
Cell Culture Techniques
Chickens
Extracellular Space - metabolism
Membrane Transport Proteins - biosynthesis
Membrane Transport Proteins - isolation & purification
Membrane Transport Proteins - pharmacology
N-glycosylation
Protein Binding
Recombinant Proteins - isolation & purification
Recombinant Proteins - metabolism
Riboflavin - metabolism
Riboflavin-binding protein
Taste - drug effects
Time Factors
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Summary:Riboflavin-binding protein (RBP) is a glycophosphoprotein found in hen eggs. We previously identified the extraordinary characteristic of RBP in reducing bitterness. For a more detailed study on the mode of action and industrial application of this characteristic, we investigated the microbial production of recombinant RBP (rRBP). We constructed a chicken RBP gene expression vector by inserting the RBP cDNA in pNCMO2, the Escherichia coli–Brevibacillus choshinensis shuttle vector. B. choshinensis HPD31 transformants produced 0.8g/l of processed and unglycosylated RBP in a soluble form in the culture supernatant. However, the expressed RBP was partially dimerized and monomeric RBP was purified by two step anion-exchange and gel-filtration chromatographies. The purified rRBP elicited bitterness reduction against quinine and caffeine, although it largely lost its riboflavin-binding ability. These results indicated that glycosylation and riboflavin-binding ability are not essential for the bitterness reduction of RBP. In addition, we assessed the usefulness of the Brevibacillus system for the expression and secretion of RBP as a new type of bitterness inhibitor.
ISSN:1046-5928
1096-0279
DOI:10.1016/j.pep.2009.12.016