Microbial production of sensory-active miraculin.
Miraculin (MCL), a tropical fruit protein, is unique in that it has taste-modifying activity to convert sourness to sweetness, though flat in taste at neutral pH. To obtain a sufficient amount of MCL to examine the mechanism involved in this sensory event at the molecular level, we transformed Aspergillus oryzae by introducing the MCL gene. Transformants were expressed and secreted a sensory-active form of MCL yielding 2 mg/L. Recombinant MCL resembled native MCL in the secondary structure and the taste-modifying activity to generate sweetness at acidic pH. Since the observed pH-sweetness relation seemed to reflect the imidazole titration curve, suggesting that histidine residues might be involved in the taste-modifying activity. H30A and H30,60A mutants were generated using the A. oryzae-mediated expression system. Both mutants found to have lost the taste-modifying activity. The result suggests that the histidine-30 residue is important for the taste-modifying activity of MCL.
Ito
Keisuke
K
Department of Applied Biological Chemistry, Graduate School of Agricultural and Life Sciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657, Japan.
Asakura
Tomiko
T
Morita
Yuji
Y
Nakajima
Ken-ichiro
K
Koizumi
Ayako
A
Shimizu-Ibuka
Akiko
A
Masuda
Katsuyoshi
K
Ishiguro
Masaji
M
Terada
Tohru
T
Maruyama
Jun-ichi
J
Kitamoto
Katsuhiko
K
Misaka
Takumi
T
Abe
Keiko
K
D001236:Aspergillus oryzae / Q000235:genetics / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D006023:Glycoproteins / Q000008:administration & dosage / Q000096:biosynthesis* / Q000737:chemistry*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D015202:Protein Engineering / Q000379:methods; D011994:Recombinant Proteins / Q000008:administration & dosage / Q000096:biosynthesis; D013549:Sweetening Agents / Q000008:administration & dosage / Q000737:chemistry* / Q000378:metabolism*; D013649:Taste / Q000187:drug effects
D006023:Glycoproteins; D011994:Recombinant Proteins; D013549:Sweetening Agents; C008533:miraculin protein, Synsepalum dulcificum