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US 5,215,963 AGranted
Solubilization and purification of the active gastrin releasing peptide receptor
AssigneeBerlex Laboratories, Inc.
InventorsRichard I. Feldman, James M. Wu, Elaina Mann, Anne Larocca, James C. Jenson
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Abstract
A procedure for the preparation of the solubilized and purified gastrin releasing peptide receptor, in an active form, from a gastrin releasing peptide receptor source such as Swiss 3T3 fibroblasts.
Claims27
A method for solubilizing the gastrin releasing peptide receptor in an active form comprising the steps of:
- (1) preparing membranes from a gastrin releasing peptide receptor source;
- (2) washing said membranes and suspending said membranes in a buffer;
- (3) solubilizing the gastrin releasing peptide receptor by incubating a culture of said membranes in a culture medium to which a detergent and a soluble cholesteryl ester has been added;
- and
- (4) adjusting the detergent concentration of said solubilized gastrin releasing peptide receptor.
2 more elements
A method for solubilizing and purifying the gastrin releasing peptide receptor in an active form comprising the steps of:
- (1) preparing membranes from a gastrin releasing peptide receptor source;
- (2) washing said membranes and suspending said membranes in a buffer;
- (3) solubilizing the gastrin releasing peptide receptor by incubating a culture of said membranes in a culture medium to which a detergent and a soluble cholesteryl ester has been added, forming a soluble extract;
- (4) precipitating the gastrin releasing peptide receptor from the soluble extract of step (3), collecting the precipitate by centrifugation and resuspending said precipitate in a buffer containing said detergent and soluble cholesteryl ester, forming a soluble extract;
- (5) applying the soluble extract of step (4) to a wheat germ agglutinin affinity column forming an eluate;
- (6) further fractionating the wheat germ agglutinin column eluate from step (5) on a gastrin releasing peptide affinity column, forming an eluate;
- and
- (7) dialyzing the gastrin releasing peptide affinity column eluate from step (6) and applying said eluate to a second gastrin releasing peptide affinity column.
5 more elements
Dependencies are inferred from the claim text.