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Description
This programme is one of a series of six which concentrate on crucial biochemical techniques, their practice and applications presented in a studio-based laboratory setting. In the programme Prof...essor Rose and Dr. Cohen complete five and begin the sixth stage of a seven stage purification of the enzyme, yeast hexokinase. The techniques involved are centrifugation, acid precipitation of protein, salting out, vacuum filtration and dialysis; these techniques are demonstrated along with other associated laboratory practice. The completion of the sixth and seventh stages is achieved in programme 2.
Metadata describing this Open University video programme
Item code: S2-1; 01
First transmission date: 25-06-1972
Published: 1972
Rights Statement:
Restrictions on use:
Duration: 00:24:31
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Producer: Roger Jones
Contributors: Norman Cohen; Stephen Rose
Publisher: BBC Open University
Keyword(s): Acid precipitation of protein; Applications; Biochemical techniques; Centrifugation; Dialysis; Enzyme purification; Practice; Salting out; Stage 5-6; Vacuum filtration
Footage description: Stephen Rose introduces the unit. He discusses the importance of experimental techniques in biochemistry. The programmes in this course will therefore feature several experimental techniques which are crucial to the field. Rose points out some of the laboratory equipment which will be used. This programme will feature techniques used to isolate the enzyme hexokinase from yeast. Rose explains why yeast was chosen as the starting material. Rose begins the process of breaking up yeast cells. He rolls the yeast out with a rolling pin to make the cell walls fragile (The yeast is then dried for several days. This is not shown.) N.R.Cohen mixes the dried yeast with a buffer to extract-the proteins. The mixture is heated to speed up the process. Stephen Rose uses the technique of centrifugation to separate the soluble from the non-soluble proteins in the mixture. He pours the mixture into test tubes, which will be centrifuged. The cell debris will be spun out. N.R.Cohen removes the tubes from the centrifuge. A heavy precipitate of cell debris has formed at the bottom. Cohen pours off the solution from the top of the test tubes. This contains proteins in solution. Cohen explains and demonstrates a method for bringing more of the proteins out of solution. The charge of the proteins is changed by altering the Ph of the solution with acetic acid. This brings proteins out of solution (ph is monitored with a ph meter). The enzyme hexokinase, however, remains in solution as the ph has not been lowered sufficiently for it to precipitate. Cohen explains and demonstrates the "salting out" process for precipitating proteins from a solution. Ammonium sulphate is the salt used. Cohen uses filtration technique to separate the precipitated proteins from the liquid. Rose adds more ammonium sulphate to the liquid fraction and explains that the precipitate would then be spun out in the centrifuge. Cohen with the result of the above. He cants off the liquid from the test tube. Cohen adds distilled water to redissolve the enzyme so that it can be further purified. Stephen Rose demonstrates and explains the process of dialysis which removes the ammonium sulphate from the solution.
Master spool number: 6LT/70231
Production number: 00521_2127
Videofinder number: 667
Available to public: no