Re: tryptic digest

L. Ericsson (ericsson@u.washington.edu)
Thu, 19 Nov 1998 09:38:57 -0800 (PST)

The best discussion I am aware of concerning "atypical" cleavages by
trypsin and other proteolytic enzymes was done by the late Borivoj Keil in
his monograph "Specificity of Proteolysis" published by Springer-Verlag
1992. Keil collected literature data on proteolytic cleavages observed by
investigators during determination of protein sequences the old fashion,
peptide by peptide. This data was accumulated in the data bank known as
LYSIS and was the basis for the monograph.

Concerning tryptic cleavages involving proline he wrote: "Proline residue
in position P'1 usually blocks trypsin action; however, the bond R-P can
be cleaved if preceded by Gln, as was observed for sequences -QR/PD,
-QR/PG, and -AGGR/PQ." (He gives 5 references) "A statistical study, made
with the help of the database LYSIS, has shown negative influences of
residues surrounding the R- and K-bonds during trypsin cleavages of a pool
of protein substrates. It illustrates in particular: the strong negative
influence of proline in position P'1 (which) is more pronounced for Lys-
than Arg-bonds, which is consistent with the role of Arg- as a stronger
substrate for trypsin than Lys-."

-Lowell Ericsson, Dept. of Biochemistry, U. of Washington, Seattle, WA

On Thu, 19 Nov 1998, Peter Hunziker wrote:

> long time ago my supervisors teached me that trypsin does never cleave the
> peptide-bond between LYS/ARG and PRO. Now, I also tell my students that
> this bond is not cleaved at all. However, in a recent LC/MS analysis we
> measured the masses of two peptides that indicated a tryptictic cleavage
> exactly at this bond (Enzyme: mod. trypsin from Promega). Since I could not
> believe it we additionally sequenced the peptides by Edman degradation and
> it turned out that trypsin has cleaved the LYS-PRO bond.
> On the C-terminal side of the cleavage site there were a number of GLY
> residues. Does anybody know if this may influences the specificity of the
> enzyme? Were my supervisors wrong and trypsin does cleave LYS/ARG-PRO bonds?
>
> best regards
> Petre
>
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