A nonapeptide was determined to have the following amino acid composition: (Lys)₂, (Gly)₂, (Phe)₂, His, Thr, Met.  The native peptide was incubated with 1-fluoro-2,4-dinitrobenzene (FDNB) and then hydrolyzed; 2,4-dinitrophenylhistidine was identified by HPLC.  When the native peptide was exposed to cyanogen bromide (CNBr), an octapeptide and free glycine were recovered.  Incubation of the native peptide with trypsin gave a pentapeptide, a tripeptide, and free Lys.  2,4-Dinitrophenyl-histidine was recovered from the pentapeptide, and 2,4-dinitrophenylphenylalanine was recovered from the tripeptide.  Digestion with the enzyme pepsin produced a dipeptide, a tripeptide, and a tetrapeptide.  The tetrapeptide was composed of (Lys)₂, Phe, and Gly.  The native sequence was determined to be:
*Recall: Pepsin cleaves N-terminal peptide bond of F, Y, W & L residues.*

• A. Gly—Phe—Lys—Lys—Gly—Thr—Met—Phe—His.
• B. His—Thr—Gly—Lys—Lys—Phe—Phe—Gly—Met.
• C. His—Thr—Phe—Gly—Lys—Lys—Phe—Met—Gly.
• D. His—Phe—Thr—Gly—Lys—Lys—Phe—Met—Gly.
• E. Met—Thr—Phe—Lys—Phe—Gly—Gly—Lys—His.

Answer: (C)

Verified step by step guidance

1. Identify the N-terminal amino acid using FDNB. The reaction with FDNB and subsequent hydrolysis indicates that histidine is at the N-terminal, as 2,4-dinitrophenylhistidine was identified.
2. Use the information from the CNBr cleavage. CNBr cleaves at methionine residues, producing an octapeptide and free glycine, indicating that glycine is at the C-terminal of the peptide.
3. Analyze the trypsin digestion results. Trypsin cleaves at the C-terminal of lysine and arginine. The presence of free lysine and the recovery of 2,4-dinitrophenylhistidine from the pentapeptide suggests that histidine is at the N-terminal of the pentapeptide.
4. Consider the pepsin digestion results. Pepsin cleaves at the N-terminal of phenylalanine, tyrosine, tryptophan, and leucine. The tetrapeptide composition of (Lys)2, Phe, and Gly suggests that phenylalanine is involved in the cleavage, helping to determine the sequence.
5. Combine all the information to deduce the sequence. The N-terminal histidine, the C-terminal glycine, and the cleavage patterns from CNBr, trypsin, and pepsin help to piece together the sequence: His—Thr—Phe—Gly—Lys—Lys—Phe—Met—Gly.