3. Approximate molecular weight for an unknown protein from gel-filtration experiment is 130 kDa. Thirty six mg of this pure protein was treated with excess of fluorodinitrobenzene. After the reaction and complete acid hydrolysis the mixture was found to contain 356 µg of dinitrobenzene derivative of methionine (free acid) and no other amino acid-dinitrobenzene derivatives. Is this protein consisting of single polypeptide chain or multiple subunits? If it consists of multiple subunits, then how many? From these data, can you calculate the molecular weight of the protein more precisely?

Which of the peptide sequences below best matches the hydropathy plot shown? 1. ILYYAGSREDHSGYLIL 2. RAFLILFMTYFLILFLI 3. LHGDQNRERDGHSQERD 4. LIFLAIFPAGSTSEDRR 5. EQSDTERNQHGALIYLI

14. A protein mixture consisting of proteins A, B, and C was subjected to various protein purification steps. First, the protein mixture was applied to an anion ion exchange column. When the column was washed with an increasing concentration gradient of chloride ions, the order of elution of the three proteins was B, then C, and finally A. Next, the protein mixture was chromatographed on a gel filtration column. The order of elution of the three proteins from this column was C, A, B. Finally, the protein mixture was passed over a chromatography column containing a Ni-NTA affinity matrix. Only protein A was retained by the column. Answer the following (i) Which protein (A, B, or C) has the highest and lowest positive charge (ii) Which protein has the highest and lowest mass (ii) Which protein has an affinity for the Ni-NTA matrix and why

The following proteins were separated by SDS-PAGE in the presence of mercaptoethanol. Sketch the relative positions of the various polypeptides on the gel. Label the positive and negative ends of the gel.Protein A: 40 kDa single polypeptideProtein B: 80 kDa protein, made up of two subunits of molecular weight 20 kDa and 60 kDa, held together by noncovalent interactionsProtein C: 200 kDa protein, made up of four identical subunits (50 kDa each) linked together by disulfide bonds