[3dem] crosslinking reagents used for successful cryo-EM samples

[Brian Gibbons]

Hi Sasha,

I found Holger Stark's GraFix method very helpful for Cryo-EM work with
transcription factor and polymerase complexes, which may be similar to your
situation.  Holger's paper covers the technique, and you can also look at
my and other publications that benefited from this technique; some links
below.  Happy to discuss further.

Good luck!
Brian

Holger's paper: https://urldefense.proofpoint.com/v2/url?u=https-3A__pubmed.ncbi.nlm.nih.gov_20887855_&d=DwIBaQ&c=-35OiAkTchMrZOngvJPOeA&r=L7-zyQ-04fFCMRqzLIOnx7H0exGZHwIQe_wMPuY600I&m=DNcmcw2ienZ_yQ624pixtXSpjjgFaZqmD6xLZnsRU9YZrY_KqbdpPTo0VRc3A0tf&s=2kWdQFZQkdFBxBbrPawQNBJRSI9MJihWzJeGVTWMQyM&e= 
- PMID: 20887855
- DOI: 10.1016/S0076-6879(10)81005-5
<https://urldefense.proofpoint.com/v2/url?u=https-3A__doi.org_10.1016_s0076-2D6879-2810-2981005-2D5&d=DwIBaQ&c=-35OiAkTchMrZOngvJPOeA&r=L7-zyQ-04fFCMRqzLIOnx7H0exGZHwIQe_wMPuY600I&m=DNcmcw2ienZ_yQ624pixtXSpjjgFaZqmD6xLZnsRU9YZrY_KqbdpPTo0VRc3A0tf&s=B8fLicxnEuHEAYzu9stkDunWm2FF-WFq8Qeyb7j9AWo&e= >
-
-
-
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Other work where this technique was applied to stabilize protein
interactions for structural work:

https://urldefense.proofpoint.com/v2/url?u=https-3A__www.pnas.org_content_109_6_1949&d=DwIBaQ&c=-35OiAkTchMrZOngvJPOeA&r=L7-zyQ-04fFCMRqzLIOnx7H0exGZHwIQe_wMPuY600I&m=DNcmcw2ienZ_yQ624pixtXSpjjgFaZqmD6xLZnsRU9YZrY_KqbdpPTo0VRc3A0tf&s=jKbp75Nznd_-_8b7WB33uTbKA08IByCB82Zl8JnDdm8&e= 
https://urldefense.proofpoint.com/v2/url?u=https-3A__doi.org_10.1073_pnas.1105266109&d=DwIBaQ&c=-35OiAkTchMrZOngvJPOeA&r=L7-zyQ-04fFCMRqzLIOnx7H0exGZHwIQe_wMPuY600I&m=DNcmcw2ienZ_yQ624pixtXSpjjgFaZqmD6xLZnsRU9YZrY_KqbdpPTo0VRc3A0tf&s=m_2VTcbo96Jfm0eW2VOlsGXlMmZ4xhgpTHatKhe2y8w&e= 
https://urldefense.proofpoint.com/v2/url?u=https-3A__www.ncbi.nlm.nih.gov_pmc_articles_PMC4039082_&d=DwIBaQ&c=-35OiAkTchMrZOngvJPOeA&r=L7-zyQ-04fFCMRqzLIOnx7H0exGZHwIQe_wMPuY600I&m=DNcmcw2ienZ_yQ624pixtXSpjjgFaZqmD6xLZnsRU9YZrY_KqbdpPTo0VRc3A0tf&s=OxqvkSEElNWpz5-i49XfosNh2AGTKJ-hxhc4p4NXFik&e=   Science. 2013 Nov 8;
342(6159): 1238724
<https://urldefense.proofpoint.com/v2/url?u=https-3A__www.ncbi.nlm.nih.gov_entrez_eutils_elink.fcgi-3Fdbfrom-3Dpubmed-26retmode-3Dref-26cmd-3Dprlinks-26id-3D24072820&d=DwIBaQ&c=-35OiAkTchMrZOngvJPOeA&r=L7-zyQ-04fFCMRqzLIOnx7H0exGZHwIQe_wMPuY600I&m=DNcmcw2ienZ_yQ624pixtXSpjjgFaZqmD6xLZnsRU9YZrY_KqbdpPTo0VRc3A0tf&s=6h7zV0kLAXdvy-X6UtpJMFalAcmNSRTvPyPO2HR6lTc&e= >
  doi: 10.1126/science.1238724
<https://urldefense.proofpoint.com/v2/url?u=https-3A__dx.doi.org_10.1126-252Fscience.1238724&d=DwIBaQ&c=-35OiAkTchMrZOngvJPOeA&r=L7-zyQ-04fFCMRqzLIOnx7H0exGZHwIQe_wMPuY600I&m=DNcmcw2ienZ_yQ624pixtXSpjjgFaZqmD6xLZnsRU9YZrY_KqbdpPTo0VRc3A0tf&s=ERkqyGKhvUQa6Hi41SW_P6zIRERnlx62rKVZAY29YY8&e= >








On Mon, Nov 22, 2021, 5:28 PM SSS Sasha <sss_jiandan at outlook.com> wrote:

> Hi all,
>
> We are working with two protein complexes that weakly interact with each
> other. We cannot isolate a stable complex using gel filtration or other
> similar approaches. We are wondering if it is possible to capture the
> complex and interaction using crosslinking reagents. We have seen the use
> of BS3 in some cryo-EM structures and wondering if you have suggestions and
> advice on other crosslinking reagents that have been successfully used for
> high resolution cryo-EM structures, and how to select the best reagent and
> condition.
>
> Thank you very much for your help,
>
> Shasha
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> 3dem at ncmir.ucsd.edu
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>
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