Methods in Enzymology Vol.102 Hormone Action, Part G: Calmodulin and Calcium-Binding Proteins

This chapter discusses the historical perspective of calmodulin purification and the strategies behind approaching the purification of calmodulin from specific tissues. The chapter discusses conventional approaches to purify calmodulin from readily available tissues, such as the rat or bovine testes, the bovine brain, outdated human red blood cells, the chicken gizzard, and the electric organ of Electrophorus electricus. The procedure involves the homogenization of the tissue in a buffered ethylenediaminetetraacetic acid (EDTA) solution, centrifugation, heat treatment, ion exchange, and gel permeation chromatography. The presence of calmodulin during the purification procedure can be assessed by a number of methods including commercially available radioimmunoassay (RIA) kits, or calmodulin-deficient cyclic nucleotide phosphodiesterase. The economical method is the differential migration of calcium-bound calmodulin during gel electrophoresis. Once purified, calmodulin can be used as an important reagent in probing a variety of systems for its mechanism of action. The protein can be used to elicit monospecific antibodies, bound to affinity resins to purify binding proteins, labeled with iodine, prepared as a photoaffinity label, or fluorescently labeled. The modification of calmodulin can result in the retention of complete properties of the native protein.

• Series: Methods in Enzymology (Book 102)
• Hardcover: 368 pages
• Publisher: Academic Press; 1 edition (October 12, 1983)
• Language: English
• ISBN-10: 0121820025
• ISBN-13: 978-0121820022
• Product Dimensions: 6 x 0.8 x 9 inches

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