1. You have a mixture of the following proteins:

 

Protein

Molecular Weight

pI

Soluble in 66% Saturated Ammonium Sulfate

Eenie

65,000

4.7

Yes

Meenie

18,000

4.4

No

Miney

30,000

8.2

No

Moe

110,000

7.9

No

Fred

85,000

7.2

Yes

 

    1. Which proteins will stick to a Diethylaminoethyl Cellulose (anion exchanger) at pH 6.5?

Eenie And Meenie.  The pH is greater than their pIs, so they will ne negatively charged.

 

    1. Which proteins will stick to a Carboxymethyl Cellulose (cation exchanger) at pH 6.5?

Miney, Moe, and Fred – same reason, opposite effect.

 

    1. What happens if, after (a) I raise the pH to 7.5?

The question was supposed to read “after  (b),” whereupon Fred would decoupe.  As I miswrote the question, the answer is “absolutely nothing.”

 

    1. In what order will the proteins migrate on an SDS-PAGE gel (farthest to least far)?

Farthest: Meenir, Miney, Eenie, Fred, Moe: Least far.

 

    1. If I add b-mercaptoethanol to the protein mixture prior to SDS-PAGE, I notice that the band for Moe disappears and two new bands appear with molecular weights of 40,000 and 70,000.  What does this tell you?

Moe is composed of two separate polypeptide chains of 40 and 70 kDal, connected by a disulfide bridge.

 

    1. In what order will the proteins elute from a gel filtration column with a molecular weight cutoff for globular proteins of 200,000?

Moe, Fred. Eenie, Miney, Meenie

 

    1. In what order will the proteins elute from a gel filtration column with a molecular weight cutoff for globular proteins of 75,000?

Moe and Fred together (at the void volume), then Eenie, then Miney, then Meenie

 

    1. In what order will the proteins appear on an Isoelectric Focusing gel, starting from the cathode end and going to the anode end.  (You might want to look up "cathode" and "anode" before answering).

Miney, Moe, Fred, Eenie, Meenie

 

  1. I cleave a protein with Cyanogen Bromide and get 2 fragments, A and B.  Sequencing of fragment B reveals that it ends (C-terminal) in homoserine lactone.  Which end of the protein does this fragment represent?

The amino-terminal end.  Since it ends at homoserine lactone (which used to be a Met), that means CNBr must have cleaved after this fragment..

 

  1. I determine the structure of a protein from a bacterium isolated from a newly-fallen meteorite.  The protein has an odd right-handed helical secondary structure, where carbonyl Oxygen atoms are hydrogen bonded to amide nitrogens on the sixth amino acid farther up the chain.  X-ray studies indicate that there are 4.7 amino acids per turn of the helix.  Given that there are currently known secondary structures named the 310 helix, the 3.613 helix, and 4,116 helix, what should I call this structure?

4,719 helix,