Dear Jiamu Du --
I want to crystallize a mAb Fab in complex with a hydrophobic
cyclic peptide. The peptide contains 1 hydrophilic residue, 2 Cys,
and 9 hydrophobic residues. The problem is that I can not dissolve
this peptide in common buffers. I think it is need to dissolve it
in some organic solvent, such as DMSO. But organic solvent is
harmful to protein.
Is there any sugestions for this situation?
This has been addressed before. If I recall quote the previous answer
properly:
" In the case of insoluble ligands, solubilize the ligand in DMSO so
it is maximally concentrated, 100mM works fine. Add enough compound
to achieve 2-3 fold excess to the protein, mix and set up. Make sure
the final DMSO concentration is ~3%, otherwise chances are it harms
the protein. If one cannot achieve a high enough stock concentration
of DMSO to be below the 3% threshold, dilute the protein in the
storage buffer to ~1mg/ml. Add compound to 2-3 fold access, incubate
and co-concentrate to the desired concentration. That way it avoids
the DMSO shock. Alternatively one could incubate the concentrated
protein with the compound solubilized in water for 24-48hr and hope
it is soluble and potent enough to get taken up by the protein and
then set up the trays."
Please note here I'm only referring to a previous e-mail from the bb.
Never tried it myself.
HTH
Kind regards.
Leo
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Chavas Leonard, Ph.D.
Research Associate
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Faculty of Life Sciences
The University of Manchester
The Michael Smith Building
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M13 9PT
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