Dear Professor, These crystals contain dihexamer insulin. That is, the same as 1XDA (insulin detemir). Therefore, the unit cell value at 100K is certainly correct. It is also confirmed by the literature that the other unit cells at 200K and 300K are hexamer insulin. The 200K and 300K space groups are not I213 but R3:H. The 200K and 300K structure determination was completed without any error when I analyzed using the R3:H space group. However, I got an error during the MR even though I used the real dihexamer structure, 1XDA (H3 double c). I can share the original data directly. Thank you very much 9 Eki 2024 Çar 16:27 tarihinde Dimitris Triandafillidis < [email protected]> şunu yazdı:

Dear Esra,

I do not believe that a transition from H3 to I213 is possible, because these two crystal forms contain different oligomeric forms of the protein in the asu. The rhombohedral form contains a hexamer (dimer in the asu), while the cubic one contains a dimer (monomer in the asu). Since these two oligomeric forms are not crystallographically related, I doubt you can induce such a rearrangement in the unit-cell just by temperature.

My suspicion is that the 100 K double c axis is due to lattice strains during freezing. Speaking from experience, insulin crystals are a bit finicky to work with, they are very fragile and these axis doubling effects are very common when the crystal is not cryoprotected well enough or not flash frozen fast enough, therefore it's more of a handling artifact. Now if all these datasets are from the same crystals and you first measured at 100 K and then increased the temperature, that would explain why the lattice strains are relieved as the crystal slowly heats up. If not, it's hard to tell without further details on the crystallization condition or collection on more crystals whether the double c axis cell at 100 K is a fluke or not.

Now, regarding your MR issue, there are a few double c insulin structures in the PDB (human, bovine or porcine) that you could use instead for starting models: - H3 double c [78.6 78.6 79.1 90 90 120 / R6 conformation] : 1XDA, 3P33, 5URU - H3 double c [80.4 80.4 72.0 90 90 120 / T3R3 conformation] : 1FU2, 1G7A, 1G7B - H3 double c [82.7 82.7 68.1 90 90 120 / T6 conformation] : 3W80 Judging from the provided lattice parameters, you're probably in the R6 conformation. Note that some of these entries have mutations and/or other chemical modifications that you'll probably need to get rid off eventually during model building - still could be helpful as starting models.

Hope this helps!

Best of luck! Dimitris ______________________________

*Dimitris P. Triandafillidis*

Hamburg Centre for Ultrafast Imaging Universität Hamburg HARBOR (Building 610) Luruper Chaussee 149, DE-22761 https://www.google.com/maps/search/Luruper+Chaussee+149,+DE-22761+Hamburg,+Germany?entry=gmail&source=g Hamburg, Germany https://www.google.com/maps/search/Luruper+Chaussee+149,+DE-22761+Hamburg,+Germany?entry=gmail&source=g

On Wed, Oct 9, 2024 at 11:21 AM John R Helliwell wrote:

...

Dear Esra, Building on Kay’s input, this paper goes part way to help your further investigation/understanding:- Preliminary study of a phase transformation in insulin crystals using synchrotron-radiation Laue diffraction https://doi.org/10.1107/S0108768188005956 doi.org https://doi.org/10.1107/S0108768188005956 https://doi.org/10.1107/S0108768188005956 https://doi.org/10.1107/S0108768188005956 It also provides various other insulin crystallography references to help you along with it. Best wishes, John

Emeritus Professor John R Helliwell DSc

On 8 Oct 2024, at 19:41, Esra A wrote:

 Dear Phenix community,

I performed a temperature shift data collection for di-hexamer insulin. I collected single crystal data at 100K, 200K, and 300K, respectively After processing the data, I noticed that the unit cell of the 100K dataset is distinct from 200K and 300K. 100K dataset = 78.2182 78.2182 79.3565 90 90 120 (assume that this is di-hexamer) 200K dataset = 78.3157 78.3157 39.6367 90 90 120 (it is hexamer) 300K dataset = 80.7109 80.7109 39.6399 90 90 120 (it is hexamer) Even if I completed the structure determination of 200K and 300K successfully, I couldn't complete the molecular replacement of the 100K dataset due to the "*The composition entered will not fit in the unit cell volume*" error message. I tried every "copies" and "mass" parameter in *automr *script. However, i got the error message every run. I can share with you the mtz and reference pdb file, if needed. Could you help me, please? Where am I making a mistake?

Thank you very much.

Sincerely,

Esra Ayan _______________________________________________ phenixbb mailing list -- [email protected] To unsubscribe send an email to [email protected] Unsubscribe: phenixbb-leave@%(host_name)s

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hello On Wed, Oct 9, 2024, 3:31 PM Kay Diederichs wrote:

Dear Dimitris and Esra,

this is interesting and important information, thanks! I wouldn't pretend that I am knowledgable about insulin; it's just that I'm interested in spacegroups and their relations. But I overlooked the fact that the gamma angle is 120° for the 100K data, which rules out my I 21 3 suggestion.

"R6 conformation" looks relevant. But to deduce information from the cell parameters, one also needs the information whether they refer to a P or I or C or R - centered cell.

Best wishes, Kay _______________________________________________ phenixbb mailing list -- [email protected] To unsubscribe send an email to [email protected] Unsubscribe: phenixbb-leave@%(host_name)s