ac5b01960_si_001.pdf (213.16 kB)
Unexpected Trypsin Cleavage at Ubiquitinated Lysines
journal contribution
posted on 2015-12-17, 09:05 authored by Meghan
C. Burke, Yan Wang, Amanda E. Lee, Emma Kimm Dixon, Carlos A. Castaneda, David Fushman, Catherine FenselauUnexpected tryptic
cleavage has been characterized at modified
K48 residues in polyubiquitins. In particular, the tryptic products
of all seven of the lysine-linked dimers of ubiquitin and of three
trimerslinear Ub–48Ub–48Ub, linear Ub–63Ub–63Ub, and
the branched trimer [Ub]2–6,48Ubhave
been analyzed. In addition to the peptide products expected under
commonly used tryptic conditions, we observe that peptides are formed
with an unexpected ε-glycinylglycinyl-Lys carboxyl terminus
when the site of linkage is Lys48. Trypsin from three different commercial
sources exhibited this aberration. Initial cleavage at R74 is proposed
in a distal ubiquitin to produce a glycinylglycinyl-lysine residue
which is bound by trypsin.