Cyanogen peptide cleavage

Cytochrome c(l-65) and -(66-104), derived by cyanogen bromide cleavage, form a stable complex in which the 65-66 peptide bond spontaneously slowly re-forms to give a protein with full biological activity.1124 ... 

Spencer, E. M. 1974. Amino acid sequence of the alanyl peptide from cyanogen bromide cleavage of bovine plasma albumin. Arch. Biochem. Biophys. 165, 80-89. 

Cyanogen bromide cleavage of the polypeptide subunit should yield 4 peptides if the subunit contains 3 methionines (Table III). Cleavage products were isolated by gel filtration through Sephadex G-75 columns and analyzed for purity with polyacrylamide gel electrophoresis in 8 M urea containing 0.1 M thioglycolate. Four peptide products were obtained (34). 

Cyanogen bromide cleaves peptides at the carboxyl side of Met. There is only one Met in the structure (see the fifth trypsin fragment), and the cyanogen bromide cleavage gives a hexapeptide consisting of the first six amino acids of this fragment. The N-terminal amino acid must be Tyr. We now know the first ten and the last two amino acids of p-endorphin. 

Figure 5-3. Mechanism of cyanogen bromide cleavage. Cyanogen bromide cleaves peptide chains at methionine residues. The reagent attacks the sulphur atom, and methylisothio-cyanate is cleaved from the resulting adduct,...

Fig. 19. Topography of the NBS cleavage of the six tyrosyl peptide links of native and

Cyanogen bromide cleavage M is 10th position, C-terminal residues are (2S,L,W) Amino-terminal residues (G,K,S,Y), tryptic peptide, ends in K Amino-terminal sequence SYGK... 

Because cyanogen bromide cleaves on the C-side of Met, any peptide containing Met must have Met as its C-terminal amino acid. The peptide that does not contain Met must be the C-terminal peptide. We know that peptide 3 is the N-terminal peptide because it contains Leu. Since it is a hexapeptide, we know that the 6th amino acid in the 13-amino acid peptide is Met. We also know that the eleventh amino acid is Met because cyanogen bromide cleavage gave the dipeptide Thr, Val. 

Figure 1. "Western blot" analysis of cyanogen bromide cleavage of CPa-1. Lanes A (- CNBr) and B (+ CNBr) were stained with coomassie blue, probed with the monoclonal antibody FAC2, or probed with avidin-peroxidase to detect biotinylated peptides.

Luo, K., Hurley, T., and Sefton, B. M. (1991) Cyanogen bromide cleavage and proteolytic peptide mapping of proteins immobilized to membranes. In Methods in Enzymology (T. Hunter and B. M. Sefton, eds.), Vol. 201, pp. 149-152. Academic Press, San Diego. 

Ten peptide Suctions from the cyanogen bromide cleavage of bacteriorhodopsion were resolved on an aminopropyl column (2 = 272nm) using a 45-min 7/1/1 - 1/7/1 chloroform/methanol/isopropylamine gradient [792]. Peak shapes were good and separation was adequate. 

Figure 6 Ideal SEC of myoglobin fragments and a mixture of synthetic peptide standards. Column same as Fig. 2. Mobile phase 50 mM KH2P04, containing 500 mM KCI and 8M urea, pH 6.5 flow rate, 0.2mL/min temperature, 26 C. (A) Elution profile of horse heart myoglobin (Mb) and its cyanogen bromide cleavage fragments (I, II, I + II, III) (B) elution profile of horse heart Mb and synthetic peptide standards 1-5 (sequences of standards are shown in Table 1) (C) plot of log,o MW versus elution time of Mb, cyanogen bromide fragments of Mb, and the five synthetic peptide standards. (From Ref. 37.)...

The use of BNPS-skatole for the selective cleavage of the tryptophanyl peptide bond has been proved useful in sequence studies. The cleavage yields so far obtained (about 50-70%) with peptides (289, 371) and proteins (119) indicate that the reagent is the best so far available for the cleavage of the tryptophanyl peptide bond and approaches in utility the cyanogen bromide cleavage of methionine peptide bonds (375). The reagent has been successfully applied to staphylococcal nuclease... 

---