consymes A. A peptide with 12 amino acids has the following amino acid composition: 2 Met, 1 Tyr, 1 Trp.2 Glu, I Lys, I Arg, 1 Thr, 1 Asn I He Reaction of the intact peptide with fluorodinitrobenzene followed by acid hydrolysis creates a derivative of lle. A specific cleavage of the interest peptide produces fragments with the following sequences: Glu-Cys-Asn-Met-Ly Met-Glu-Thr-Arg-Trp He-Tyr (Questions 1-5 1. Which reagent was used for the specific cleavage? a) chymotrypsin b) trypsin c) V8 protease d) cyanogen bromide Which amino acids would be released when the intact peptide was treated first with V8 protease followed by treatment with cyanogen bromide? a) Glu and Met b) Glu and Lys c) Met and Lys d) Glu, Met, and Lys 3. Which treatment would result in the release of Lys and Arg from the intact peptide? a) trypsin b) trypsin followed by dansy!chloride c) trypsin followed by carbox peptidase d) trypsin followed by mild acid If this intact peptide is sequenced using the Edman degradation, which step will be part of the procedure? a) The Edman reagent will react with all 12 amino acids simultaneously. b) Lithium borohydride will react with an a-carboxyl group. c) Phenylisothlocyanate will react with an a-amino group. d) Strong acid will be used to cleave off one modified amino acid __ If this peptide is normally part of a multimeric protein composed of four identical subunits, what procedure might be needed prior to performing the Edman degradation? a) The four subunits should be separated and sequenced individually b) Two specific cleavages should be done to create two sets of fragments. c) Peptide bonds should be broken using hydrazine. d) Disulfide bonds should be reduced with mercaptoethanol.

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consymes A. A peptide with 12 amino acids has the following amino acid composition: 2 Met, 1 Tyr, 1 Trp.2 Glu, I Lys, I Arg, 1 Thr, 1 Asn I He Reaction of the intact peptide with fluorodinitrobenzene followed by acid hydrolysis creates a derivative of lle. A specific cleavage of the interest peptide produces fragments with the following sequences: Glu-Cys-Asn-Met-Ly Met-Glu-Thr-Arg-Trp He-Tyr (Questions 1-5 1. Which reagent was used for the specific cleavage? a) chymotrypsin b) trypsin c) V8 protease d) cyanogen bromide Which amino acids would be released when the intact peptide was treated first with V8 protease followed by treatment with cyanogen bromide? a) Glu and Met b) Glu and Lys c) Met and Lys d) Glu, Met, and Lys 3. Which treatment would result in the release of Lys and Arg from the intact peptide? a) trypsin b) trypsin followed by dansy!chloride c) trypsin followed by carbox peptidase d) trypsin followed by mild acid If this intact peptide is sequenced using the Edman degradation, which step will be part of the procedure? a) The Edman reagent will react with all 12 amino acids simultaneously. b) Lithium borohydride will react with an a-carboxyl group. c) Phenylisothlocyanate will react with an a-amino group. d) Strong acid will be used to cleave off one modified amino acid __ If this peptide is normally part of a multimeric protein composed of four identical subunits, what procedure might be needed prior to performing the Edman degradation? a) The four subunits should be separated and sequenced individually b) Two specific cleavages should be done to create two sets of fragments. c) Peptide bonds should be broken using hydrazine. d) Disulfide bonds should be reduced with mercaptoethanol.

Answer 1

1. The specific cleavage reagent used is c) V8 protease. V8 protease specifically cleaves at the carboxyl side of aspartic acid (Asp) and asparagine (Asn) residues. 2. When the intact peptide is treated first with V8 protease followed by treatment with cyanogen bromide, the released amino acids would be b) Glu and Lys. V8 protease cleaves at aspartic acid and asparagine residues, releasing Glu and Lys. Cyanogen bromide cleaves at methionine residues, releasing Met and Lys. 3. The treatment that would result in the release of Lys and Arg from the intact peptide is a) trypsin. Trypsin cleaves at the carboxyl side of lysine (Lys) and arginine (Arg) residues. 4. If this intact peptide is sequenced using the Edman degradation, the step that will be part of the procedure is d) Strong acid will be used to cleave off one modified amino acid. In Edman degradation, the N-terminal amino acid is labeled with phenylisothiocyanate (PITC) and then cleaved off using strong acid. 5. If this peptide is normally part of a multimeric protein composed of four identical subunits, the procedure that might be needed prior to performing the Edman degradation is d) Disulfide bonds should be reduced with mercaptoethanol. Reducing disulfide bonds with mercaptoethanol will break the disulfide bridges between the subunits, allowing for individual subunit sequencing.

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