Affinity Techniques - Enzyme Purification: Part B by Nathan P. Kaplan, Nathan P. Colowick, William B. Jakoby,

By Nathan P. Kaplan, Nathan P. Colowick, William B. Jakoby, Meir Wilchek

The significantly acclaimed laboratory typical, Methods in Enzymology, is among the such a lot hugely revered guides within the box of biochemistry. due to the fact 1955, every one quantity has been eagerly awaited, usually consulted, and praised through researchers and reviewers alike. The sequence comprises a lot fabric nonetheless appropriate at the present time - actually an important book for researchers in all fields of lifestyles sciences

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Extra info for Affinity Techniques - Enzyme Purification: Part B

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Once dissolved in water it hydrolyzes rapidly (10-15 minutes at pH 4). ~J. K.

The mixture is gently stirred at 50 ° for 2 hours. 0 containing 1 M NaCl, and 500 ml of 1 M NaC1 at 50 °. The coupled sulfanilamide is 550 /zmoles per gram of dry gel according to the nitrogen content as determined by the Kjeldahl method. 3 M sodium carbonate at pH 10 containing 200 mg of protein. The suspension is gently stirred with a magnetic stirrer at room temperature for 2 hours. 5 M NaC1 in which the gel is stored at 5 °. In the case of soybean trypsin inhibitor, the gel contains 12 ~moles per gram of dry conjugate, and the gel is able to adsorb a stoichiometric amount of trypsin which, in a subsequent step, can be quantitatively desorbed.

S may be considered as a chromatographic measure of affinity. S can be decreased by including in the eluent a substance D that will compete with A for the adsorption sites. ~. kr/(1 + kCA + k'CD) (3) At high concentration of the displacer substance, D, S approaches zero and A will be eluted at or very close to V~, which is the volume determined by molecular sieving in the absence of adsorption. It is also possible to choose k'CD so that kCA ~ 1 + k'CD. Under these conditions, S will be independent of the concentration of A, and the latter substance will travel through the bed as a symmetrical zone with the relative retention due to adsorption equal to K,,.

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