The availability of single proteins in highly purified form is a major step for their characterisation. In the case of wine proteins, many procedures have been used, allowing to purify specific components, to be used for biochemical studies or for antibodies production. The methods described to date, however, seem to be rather time-consuming, cumbersome, and do not allow to obtain the simultaneous purification of more than one or few components. Here we describe an electrophoretic preparative procedure which allows the simultaneous and complete purification of most of the protein bands present in wine in only one step. In this system, the proteins of the sample, after being electrophoretically separated in a cylindrical gel, are eluted from its bottom trough a capillary in which the electroendosmotic buffer flow generated during electrophoresis is channelled. The proteins are then collected as single components as they elute from the capillary. Two different separation systems were used for the purification of wine proteins: one in native (non-denaturing) conditions and the other in the presence of Na-dodecyl sulphate (SDS). In both cases, several fractions containing only one electrophoretic band were obtained, indicating the effectiveness of the system in the purification of nearly all the wine proteins within a single experiment. The degree of purity and the amounts of the protein components obtainable in this way warrant the possibility of their fine (bio)chemical characterisation and also to produce antibodies with high specificity. In conclusion, the results here presented represent a major methodological improvement for the study of wine proteins in connection with their effects in winemaking.

One-sptep purification of nearly all the proteins from wine by electroendosmotic preparative electrophoresis

VINCENZI, SIMONE;CURIONI, ANDREA
2003

Abstract

The availability of single proteins in highly purified form is a major step for their characterisation. In the case of wine proteins, many procedures have been used, allowing to purify specific components, to be used for biochemical studies or for antibodies production. The methods described to date, however, seem to be rather time-consuming, cumbersome, and do not allow to obtain the simultaneous purification of more than one or few components. Here we describe an electrophoretic preparative procedure which allows the simultaneous and complete purification of most of the protein bands present in wine in only one step. In this system, the proteins of the sample, after being electrophoretically separated in a cylindrical gel, are eluted from its bottom trough a capillary in which the electroendosmotic buffer flow generated during electrophoresis is channelled. The proteins are then collected as single components as they elute from the capillary. Two different separation systems were used for the purification of wine proteins: one in native (non-denaturing) conditions and the other in the presence of Na-dodecyl sulphate (SDS). In both cases, several fractions containing only one electrophoretic band were obtained, indicating the effectiveness of the system in the purification of nearly all the wine proteins within a single experiment. The degree of purity and the amounts of the protein components obtainable in this way warrant the possibility of their fine (bio)chemical characterisation and also to produce antibodies with high specificity. In conclusion, the results here presented represent a major methodological improvement for the study of wine proteins in connection with their effects in winemaking.
Oenologie 2003
9782743006495
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/11577/1340252
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