Crystallisation of Miraculin - A Taste Modifying Glycoprotein from the Fruits of Richadella Dulcifica

93 Pages Posted: 10 Jul 2017

Date Written: December 10, 1997

Abstract

The crystallisation of miraculin - a glycoprotein was studied. The protein was extracted and purified to homogeneity from the pulp of the fruits of Richadella dulcifica using a three-step chromatographic process involving Hydrophobic Interaction Chromatography, Ion Exchange Chromatography, and Gel Filtration Chromatography respectively at ordinary temperatures. The activity of the protein was ascertained using psychometric analysis: 100g/ml of the lyophilised protein provoked sweetness with 0.02M citric acid. Sodium Dodecyl Sulphate-Polyacrylamide Gel Electrophoresis was used to determine the purity of the protein.

Experimental conditions used in 80 different crystallisation trials were chosen for the initial screening experiments. Polymorphism in the resulting precipitated, micro-crystalline, or crystalline protein samples was identified and rated numerically on a rough but quantitative scale. Correlation between these estimates of crystal quality and all the 80 different crystallisation trials were identified.

Twelve out of the 80 trials were selected for further crystallisation work by changing the concentrations of salts and precipitants and the addition of detergents until favourable conditions were achieved for the crystallisation of the protein.

Orthorhombic crystal forms of the protein with sizes ranging from 0.2mm to 0.4mm were obtained using the hanging drop technique of the vapour diffusion method from 30% PEG 600, 0.1M LiCl, 0.1M CaCl2, 0.1M HEPES buffer, 0.02M 1-O-nOctyl-D-glucopyranoside (BOG) and miraculin at a concentration of 20mg/ml. The crystalline miraculin gave a band of 44kd on SDS-PAGE

Keywords: Crystallization; Glycoprotein, Miraculin, Polymorphism

Suggested Citation

Yawson, Robert M., Crystallisation of Miraculin - A Taste Modifying Glycoprotein from the Fruits of Richadella Dulcifica (December 10, 1997). Available at SSRN: https://ssrn.com/abstract=2793819 or http://dx.doi.org/10.2139/ssrn.2793819

Robert M. Yawson (Contact Author)

Quinnipiac University ( email )

275 Mt. Carmel Avenue, SB-DNF
School of Business
Hamden, CT 06518
United States
2035825023 (Phone)

HOME PAGE: http://www.qu.edu/student-resources/directory/staff.69501.html

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