Dear all, I am trying to refine N-linked carbohydrates with REFMAC5 (CCP4 version 6.0.2) - however - it seems as if I am unable to get the stereochemistry right (when I tried to deposit the structure to PDB - they complaied about it). My protein comes from D. melanogaster and I can see density for the Asn-linked NAG (with a core alpha-1,6-fucose), the second NAG, and three mannoses. This is the header I have been using (I am not a 100% convinced it is correct - maybe someone could verify this please?):
LINK C1 NAG A 501 ND2 ASN A 105 NAG-ASN LINK O4 NAG A 501 C1 NAG A 502 BETA1-4 LINK O4 NAG A 502 C1 BMA A 503 BETA1-4 LINK O6 BMA A 503 C1 MAN A 504 ALPHA1-6 LINK O3 BMA A 503 C1 MAN A 505 ALPHA1-3 LINK O6 NAG A 501 C1 FUC A 506 ALPHA1-6 MODRES NAG A 501 NAG-b-D RENAME MODRES NAG A 502 NAG-b-D RENAME MODRES BMA A 503 MAN-b-D RENAME MODRES MAN A 504 MAN-a-D RENAME MODRES MAN A 505 MAN-a-D RENAME MODRES FUC A 506 FUC-a-L RENAME (1) I'd really appreciate if someone could confirm that I am using the correct links (i.e. the correct sugars (the protein is exressed in fly cells) and how they are linked). (2) Am I using the correct header for handling the sugars - and if not - what should I include/use instead? (3) Does someone have a working cif-file which includes all the necessary restraints for those sugars including the links, which I could use as a library for REFMAC? PLEASE!!!! (4) I am a bit confused about the naming convention of mannoses: MAN-b-D (according HICUP) has the residue code BMA, whereas MAN-a-D has MAN. However, it seems that in the REFMAC dictionary MAN seems to be used for both and there does not seem to be a MAN-a-D.cif file either. Can anyone please comment on this? THANK YOU SOOO MUCH, Petra -- Petra Verdino, PhD Research Associate The Scripps Research Institute, BCC206 10550 North Torrey Pines Road CA 92037 La Jolla, USA tel:1-858-784-2294 fax:1-858-784-2980
