Hi all,
I'm currently trying to crystallise a two domain protein which
contains several structurally important disulfides. We have a high
resolution structure of one domain (~1.4 A resolution), which reveals
quite a few solvent-exposed lysines, some of which are involved in
crystal-contacts.
The two-domain construct also crystallises, but the only crystals
obtained after extensive optimisation are stacks of thin-plates that
show poor diffraction (multiple lattices, streaky spots) to around 3 A.
I would like to attempt modification of the lysine residues by
reductive methylation or cyclic pentylation (in the hope of improving
morphology and/or diffraction), but I am worried that the reducing
conditions required for the reaction (due to the presence of the
dimethylaminoborane complex) will disrupt the native disulfides and
result in protein aggregation or denaturation. Does anyone have any
experience with reductive methylation of disulfided proteins, or know
of any references describing the same?
Thanks in advance,
Oliver Clarke.
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