Question

(a) In what order would the amino acids Arg, His, and Leu be

eluted from a carboxymethyl column at pH 6? (b) In what order

would Glu, Lys, and Val be eluted from a diethylaminoethyl

column at pH 8?

Answer 1

a)

• At ph 6 ,pka of his is 6.09,ie His side chain is 50% charged( +0.50).similarly Arg(+1),and Leu is (0).
• The C -terminal Pka is 2.2 and so it is -1 at ph 6 .
• The N-terminal Pka is 9.4 and so it is +1 at ph 6.
• carboxy methy matrix is the most commonly used cation exchanger. The molecules with posive charge will stick to the negatively charged coloum first.so the order will be Leu(0)-His(+0.5)-Arg( +1)

​b)

• At Ph 8 ,Glu(-1), Val(0) ,Lys(+1) .
• The C-terminal Pka is 2.2 so it is -1 at ph 8
• The N-terminal Pka is 9.4 so it is +1 at ph 8.
• Diethylaminoethyl is an anionic coloumn ,so the negatively charged molecules will stick to the anion exchanger and come off later .so the order would be LYS(+1)-VAl(0)- Glu(-1).