Forthe life of me, I can not figure out this question. Thanks inadvance! Iam try

Question

Forthe life of me, I can not figure out this question. Thanks inadvance!

Iam trying to purify a particular protein (Protein C) from anextract of red blood cell

proteins from Quatchi. I have gone through differentpurification steps, and currently

have a buffered solution of pH = 7.5 containing two other proteinsalong with Protein C.

Theamino acid compositions of these proteins are givenbelow.

Protein C: 25% Asn, 20% Gly, 10% Asp, 20% Ser, 20% Arg, 5%Tyr

Protein R: 30% Gln, 25% Glu, 2% Phe, 20% Lys, 13% Ser, 10%Cys

Protein X: 25% Ala, 20% Gly, 20% Ser, 10% Ile, 10% Val, 5%Asn, 5% Gln, 5% Pro

Allthree proteins are similar in size and pI, and I don’t haveany antibodies available to

me. Assuming that sasquatch proteins have the same molecularmakeup and properties as

human proteins,

a.What specific fractionation procedure should I use to purifyProtein C from this

mixture?

b.Which protein would have the greatest A280 value?

Explanation / Answer

      a.) The specificfractionation procedure that can be used to purify protein C fromthe given mixture is ionexchange           chromatography.           As this protein has Agrinine which is a positively charged aminoacid, this protein can exhibit a net positive charge           andthis can be sused to separate the protein from the othermixture.           Evenprotein R has a positively charged amino acid (Lysine) but, itspositive charge can be neutralised by the amino           acidglutamate which is negatively charged.           Hencethis protein does not exhibit any charge.           Theother protein; protein X does not have any of the charged aminoacid, hence it does not exhibit charge.           Therefore,this will not be an obstruction in the separation.          Therefore,ion exchange chromatography can be used to separate protein C fromthe given mixture.           b.)   Theabsorption of a protein is due to the presence of aromatic aminoacids in it.               Basingon this fact, the protein that shows maximum absorption at280 is protein C.              This is because, only this protein has the aromatic aminoacid in the from of Tyrosine and other proteins does not              have any of the aromatic amino acid.

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