Output Files

[Ivan S. Podkorytov]

Dear Woonghee,

May I ask you several questions about format of output file

protein_backbone_assignment.txt file ?

What is the meaning of 0.000 and 999.00 values of chemical shifts?

For example, the  output file

http://i-pine.nmrfam.wisc.edu/files/protein_backbone_assignment.txt

contains 0.0 for H and N for both PRO-37 and PRO-38. Do the zero values mean that

the corresponding peaks are absent in HSQC because prolines do not have HN?

Does any other 0.0 ppm value in the file means that the corresponding peak is absent?

The same file contains 999.00 for both CA and CB of PRO-37 in contrast to  66.00 and  32.80 for PRO-38. What do the two 999.00 mean?

In the same file, MET-01 has all P(H,N) = 0.0 and also P(no_assignment) = 0.0.

In contrast,  GLU-24 and GLY-53 also have all P(H,N) = 0.0 but P(no_assignment) = 1.0.

I would expect P(no_assignment) = 1.0 for N-terminal MET. Why all probabilities (including P(no_assignment)) are zero for MET-01?

Thank you and best wishes,

Ivan.

***************************************

Ivan S. Podkorytov
Researcher
Biomolecular NMR Laboratory
St. Petersburg State University
Botanicheskaya 17
Staryi Peterhof
Saint-Petersburg
Russia

***************************************

[Lee, Woonghee]

Dear Ivan,

 

What is the meaning of 0.000 and 999.00 values of chemical shifts?

 

In the protein_backbone_assignment.txt, chemical shifts 0 and 999 both means no assignment.

The probability that you see in that file indicates the probability for the generalized spin system. In P38 case, that particular spin system cannot be assigned because you don’t have a peak of N-HSQC for that residue. However, CA and CB can be assigned by evolved magnetization transfer from the next residue which is D39 in CBCA(CO)NH, HNCACB, etc. Therefore, you see legit probabilities in the sidechain table file.

 

Also, chemical shift 0 may mean really the 0 ppm in side chain assignments because proton’s can go that below.

 

M1 seems to be ignored because it cannot be crossvalidated because it does not have a previous residue.

 

Woonghee

 

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[Ivan S. Podkorytov]

Dear Woonghee,

Thank you for the I-PINE server. It really helps me in my work.

May I ask you to clarify some details concerning output files?

I obtained the following results:
http://zavot.nmrfam.wisc.edu/upload/script/users/210120_092413_642380/results/
The file sparky_hnca-10.txt contains the following lines with R66CA

R66H-R66CA-R66N      7.608     56.931    120.899
I67H-R66CA-I67N      8.232     56.927    122.230

Basin on these data I would expect to find an averaged value
(56.931 + 56.927)/2 = 56.929 that is ca. 56.93 for R66CA shift in the output file
protein_backbone_assignment.txt. Instead, this file contains 56.90.
Why does the actual number 56.90 differ from the averaged value 56.93?
How 56.90 was obtained?

Thank you very much and best wishes,

Ivan.



***************************************
Ivan S. Podkorytov
Researcher
Biomolecular NMR Laboratory
St. Petersburg State University
Botanicheskaya 17
Staryi Peterhof
Saint-Petersburg
Russia
***************************************

[Lee, Woonghee]

Dear Ivan,

 

I-PINE does not average chemical shifts. It changes chemical shifts to integer numbers and undergoes assignment procedure. When it is close to the end, it finds relevant peaks in each experiment and gives assignment labels. Basically, the assignment table itself are 0.1ppm and 0.01ppm resolution for heavy atoms and protons respectively because it has changed to integer numbers, however, it tries to bring more precise numbers using peak lists. If it cannot find unique one, it will just stay the same. 56.90, 56.931 are 56.927 are not really different values in spectroscopic perspective considering the digital resolution.

 

Best,

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