Using HNCA/HNCO Spectra

If you are using HNCA, HN(CO)CA, HN(CA)CO and HNCO spectra for your assignment the easiest thing is to use the Protein Sequence Assignment function in a mannner very similar to that described above. When using the CBCANH/CBCA(CO)NH spectra the Cα and Cβ resonances are matched. When using the HNCA and HNCO based spectra, it is the Cα and CO resonances which are matched. Within the Protein Sequence Assignment panel you will have to set both the HNCA and HN(CA)CO as your query spectra. In these spectra you will need to assign the Cα and CO resonances belonging to the query NH group. The matched spectra in which you look for your matches will be the HN(CO)CA and HNCO spectra. One difficulty that arises when using the HNCA/HNCO spectra is that the CO resonances have chemical shifts between about 170 and 180 ppm. So while the Cα and Cβ resonances occur at similar ppm values (about 75-15 ppm) and it is easy to look at these in the same window, this is not so easy if you want to inspect the Cα and CO resonances at the same time. To get around this problem, you can simply tick the Focus 13C Matches option in the Settings tab of the Protein Sequence Assignment pop-up.

Link Sequential Spin Systems pop-up

Analysis will then automatically add horizontal separators to your windows and focus the window sections on all the relevant peaks. This means that you can look at the HNCA and HNCO spectra in the same window rather than having to juggle additional windows or do lots of scrolling.

Matching peaks in HNCA/HNCO-based spectra

Now you can select a spin system in the Protein Sequence Assignment panel, and match it in the normal way.