Question

6. A series of proteins of known mass were running over a Sephacryl S300 column.

A. Determine the molecular mass of the unknown protein.

B. Give a plausible explanation for the strange behavior of ferritin’s elution from the column. How would you test your idea?

Protein	Mr (kDa)	Ve (mL)
Blue dextran	1000	85
Lysozyme	14	200
Chymotrypsinogin	25	190
Ovalbumin	45	170
BSA	65	150
Aldolase	150	125
Urease	500	90
Ferritin	700	92
Ovomucoid	28	160
unknown	??	130

Answer 1

Protein ellution from the column occurs according to the molecular weight of the protein. the more the molecular weight the slower is its ellution from the column.

The order of ellution according to molecular weights should be lysozyme, chymotrypsinogen, ovomucoid, ovalbumin, BSA, aldolase, urease, ferritin and blue dextran.

The order of ellusion according to Ve is lysozyme, chymotrypsinogen, ovomucoid, unknown, BSA, aldolase, ferritin, urease,Blue dextran.

A) Molecular mass of the unknown protein may be before aldolase so its molecular weight may be less than aldolase.

B) According to molecular weight ferritin should be elluded before blue dextran but is elluded before urease.