2009_Book_FoodChemistry

1.4 Proteins 43
(1.84)
The C-terminal amino acids can be removed
enzymatically by carboxypeptidase A which
preferentially cleaves amino acids with aromatic
and large aliphatic side chains, carboxypeptidase
B which preferentially cleaves lysine,
arginine and amino acids with neutral side chains
or carboxypeptidase C which cleaves with less
specificity but cleaves proline.
1.4.1.3 Partial Hydrolysis
Longer peptide chains are usually fragmented.
The fragments are then separated and analyzed
individually for amino acid sequences. Selective
enzymatic cleavage of peptide bonds is
accomplished primarily with trypsin, which
cleaves exclusively Lys-X- and Arg-X-bonds,
and chymotrypsin, which cleaves peptide bonds
with less specificity (Tyr-X, Phe-X, Trp-X and
Leu-X). The enzymatic attack can be influenced
by modification of the protein. For example,
acylation of the ε-amino group of lysine limits
tryptic hydrolysis to Arg-X (cf. 1.4.4.1.3
and 1.4.4.1.4), whereas substitution of the SHgroup
of a cysteine residue with an aminoethyl
group introduces a new cleavage position
for trypsin into the molecule “pseudolysine
residue”):
(1.85)
Also suited for the specific enzymatic hydrolysis
of peptide chains is the endoproteinase Glu-C
from Staphylococcus aureus V8. It cleaves Glu-
X bonds (ammonium carbonate buffer pH 7.8 or
ammonium acetate buffer pH 4.0) as well as Glu-
X plus Asp-X bonds (phosphate buffer pH 7.8).
The most important chemical method for selective
cleavage uses cyanogen bromide (BrCN) to
attack Met-X-linkages (Reaction 1.86).
Hydrolysis of proteins with strong acids reveals
a difference in the rates of hydrolysis of peptide
bonds depending on the adjacent amino acid side
chain. Bonds involving amino groups of serine
and threonine are particularly susceptible to hydrolysis.
This effect is due to