Hua Peter Sun and S. Radaev wrote a paper a few years ago where they showed by data mining that the crystallization conditions of complexes heavily favor PEG rather than salt conditions, so you are not alone.
Radaev and Sun, Crystallization of protein-protein complexes J. Appl. Cryst. (2002). 35, 674-676 You could try microseeding *into random screens* using the crystals that you have to make a seed stock. See Acta Cryst. (2007). D63, 550–554 and http://www.douglas.co.uk/mms.htm. This approach has sometimes worked before for complexes, eg seeding crystals of an antigen-Fab complex with crystals of just the Fab. You can use this approach by hand or with a robot (spin the seed stock if you us a non-contact robot). One problem is that you would normally put quite a lot of Amm Sulf into your wells because you would suspend the seed crystals in the reservoir solution where the crystals used to make the seed stock were found. The conventional volumes to use are 0.3 protein + 0.2 reservoir solution + 0.1 seed stock, so typically 1/3 of the precipitant would be Amm Sulf. My colleagues and I recently submitted a manuscript to Crystal Growth and Design where we look in great detail at suspending seed crystals in other precipitants (instead of Amm Sul in this case). We found that this normally works, eg you can usually make a seed stock with PEG instead of say salt conditions. You can predict whether the PEG (or any other solution) will work for suspension by incubating the crystals in PEG for 24 hours. If the crystals look unchanged after that, you can almost certainly make a seed stock from the solution in question. (If the crystals dissolve, shatter or crack, you MAY be able to make a seed stock with the solution.) We used 100% PEG600, although in restrospect I think 30% - 50% PEG 3000 might be better. I can send you a preprint, but the gist of the relevant part is above. For a review of more radical approaches, see Bing Xiao et al., Optimizing Protein Complexes for Crystal Growth. Crystal Growth & Design, Vol. 7, No. 11, 2007 2215 I hope that helps Patrick On Sun, Feb 27, 2011 at 2:24 AM, Hua Yuan <foxso...@gmail.com> wrote: > Dear CCP4 community members, > > I've been trying to crystallize a protein complex that's very sensitive to > ionic strength, i.e., lower salt (~0.3M) will cause precipitation of the > complex but higher salt (~0.5 M) breaks the complex apart. The interaction > that holds the complex is probably mainly ionic type. > The crystals I got so far has only one component of the complex from which > all the crystallization conditions have high salt such as 2M Ammonium > Sulfate in them. Besides repeatly screening many crystallization > conditions, I was wondering whether is any way to work around this problem. > Your suggestions would be greatly appreciated! > > Thanks, > > Hua > > > -- patr...@douglas.co.uk Douglas Instruments Ltd. DouglasHouse, EastGarston, Hungerford, Berkshire, RG177HD, UK Directors: Peter Baldock, Patrick Shaw Stewart http://www.douglas.co.uk Tel: 44 (0) 148-864-9090 US toll-free 1-877-225-2034 Regd. England 2177994, VAT Reg. GB 480 7371 36
