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Assign Contributions to NOE Peaks Using Structures and Chemical Shifts
This popup window is designed to assist in the latter stages of assigning NOE and other through-space spectra that are used in the generation of macromolecular structures. The system uses the matching of chemical shifts to peak positions, and a structure if available, to suggest assignments for peaks. Guiding structures are used to eliminate assignment possibilities where the resonances would be too far apart in space to give a contribution to the signal. Such structures may include comparative (homology) models or unfinalised structures from part way though the structure generation process, for example as produced by ARIA or CYANA programs.
User Operation
To operate this tool the user selects a NOESY or other though-space peak list that will be assigned. Various options may be chosen, including how assignment possibilities are displayed on a graphical structure view and what the tolerances are for considering an assignment possibility. The distance tolerance is the maximum allowable separation between potential peak dimension assignments (connected by a through-space transfer) if a guide structure is present. The F1, F2 etc. dimensional isotope tolerances are the maximum allowed chemical shift differences for matching a peak position to a resonance of known chemical shift. The “Navigation Windows” table allows the user to specify which spectrum windows will be used for navigation; to show a selected peak’s location. This table displays all windows that the peak list may be viewed in, and the “Use?” column may be toggled to activate a window for navigation.
Reciprocal Return Peaks
The active windows can not only display the location of a selected peak, but also a series of strips (window sub-divisions) that relate to any reciprocal, symmetry related (return) peaks. This is useful for the resolution of assignment ambiguity. If a peak is thought to be due to an NOE from resonance A to resonance B a reciprocal NOE (from B to A) will usually be present to confirm the assignment possibility, assuming the return peak would be visible in the spectrum. If strips for return peaks are required the user should select the “Strip Return Peaks” option at the top of the settings tab.
Peak Assignments
In the “Peak Assignments” tab the top table lists all of the peaks that are present in the selected peak list. Clicking on a peak row in this table will cause the lower table to be filled with the assignment possibilities for the peak. If navigation windows and a guide structure display is active selecting a peak in this way will also navigate to the appropriate spot in the spectrum and show structural connections.
Peak assignment possibilities in the lower table are, by default, displayed in a ranked order according to how close the peak position is to the chemical shifts and how distant in space the relevant atoms are in any guide structure. The chemical shift differences are weighted by the per-dimension tolerances so that values from dissimilar isotopes may be combined. A peak assignment is made by selecting the specific row for the resonances you wish to assign to the selected peak and clicking [Assign Selected]. An individual dimension assignment may be removed by clicking on the appropriate cell in the “Possible Assignments” table and clicking [Delete Assignment], or all assignments for the peak may be cleared with [Delete All Assignments].
The [Predict Peaks] function is designed to show you where the listed assignment possibilities would make peaks in the spectrum, given their recorded chemical shifts. This option makes synthetic peak crosses to show where on the spectrum the resonance intersections would be, and if possible what structural distance they represent. From these the user may be able to make a better judgement of which possibilities are likely to cause a peak. Such synthetic peaks are made in an entirely separate peak list to the main one, and their list may be deleted at any time without adverse effect.
Caveats and Tips
The “Strip Return Peaks” option will override any existing strips within the selected spectrum windows. If the windows should not be affected in this way the option should be switched off.
The “Tol. Weighted Combined Delta” value in the lower table is a weighted chemical shift distance (structure distance is not included) that can be used an assignment criterion, as long as your shift tolerances are set appropriately. The actual calculation is to take the shift difference for each dimension and divide by the tolerance for that dimension (normalise to be <= 1.0). All dimensions are then combined by taking the square root of the sum of squares, i.e. to give a “shift distance”. The maximum value will be the square root of the number of dimensions.
It should be noted that the [Assign Whole Peak List] will assign all of the possibilities, as listed in the lower table, for all the peaks in the entire peak list. Hence, when using this function it is recommended that suitably narrow distance and chemical shift tolerances are set.
The chemical shift list that is used for matching resonances to peak positions is the one set on the experiment record for the selected peak list.
The experiment record of a peak list must be set as a through-space type (e.g. NOESY) for it to be considered.
This system will work with non-NOESY though-space experiments such as spin diffusion types used for solid-state samples. However, at present assignment contribution fractions are only calculated using the Intensity^(1/6) summation appropriate for NOE experiments.
This system takes no account of missing assignments; resonances that may cause a peak but do not have a known chemical shift.
Sets the parameters required for matching peaks to NOE assignments, including which peak list and structure to use
NOE Peak List: Selects which NOE peak list will be considered for the matching of close resonances; in terms of both shift and distance)
Structure Display: Selects which kind of assignment possibilities, for a peak, to display on a graphical structure display
Structure: Selects which structure ensemble will be used for calculating atomic distances within the graphical structure display
Aliased Possible: Sets whether the peaks could be caused by aliased resonances; not at their real ppm value, but a whole number of sweep widths away
Focus Structure: Sets whether to rotate the graphical structure display to focus on the current assignment possibilities
Mark Peak: Sets whether to mark the selected peak position, as displayed in the selected navigation windows
8.0: Sets the upper limit for structural distance, below which pairs of close atom sites may be considered for assignment
A table of the peaks that NOE assignments are considered for, and their structure & shift filtered assignment possibilities
Table 2 | |
# | The serial number of the peak in its peak list |
Position | The position of the peak in all of its dimensions |
Assignment | The current resonance assignment annotation of the peak |
Details | A textual description of the peak; can be set from the main peak tables, e.g. to mark violated peaks |
Height | The height intensity of the peak; usually from the spectrum data matrix,but possibly set manually |
Volume | The volume integral of the peak; calculated using the method stated in the peak list table |
Previous Peak: Selects the previous peak in the table and display NOE assignment possibilities; causes navigation within spectrum windows if this is set
Selected Peak: Selects a peak in the table based upon the peak currently selected in spectrum windows; uses the first if many are selected
Next Peak: Selects the next peak in the table and display NOE assignment possibilities; causes navigation within spectrum windows if this is set
Find Reciprocal Peaks: Display strips for possible (symmetry related) NOE reciprocating return peaks, e.g. for an A-B peak look for possible B-A peaks.
Table 3 | |
# | Documentation missing |
Reson F1 | Documentation missing |
Reson F2 | Documentation missing |
Reson F3 | Documentation missing |
Reson F4 | Documentation missing |
Dist. | Documentation missing |
Tol. Weighted Combined Δ | Documentation missing |
ΔF1 | Documentation missing |
ΔF2 | Documentation missing |
ΔF3 | Documentation missing |
ΔF4 | Documentation missing |
Contrib Frac.Shift F1 | Documentation missing |
Shift F2 | Documentation missing |
Shift F3 | Documentation missing |
Shift F4 | Documentation missing |
Assign Selected: Assign the peak to the individual resonance+dimension possibility selected in the above table
Delete Assignment: Remove the assignment of the individual resonance+dimension possibility selected in the above table
Delete All Assignments: Delete all resonance assignment for all dimensions of the selected peak
Predict Peaks: Make simulated peaks (in a separate, temporary peak list) at the locations of the displayed resonance intersections
Assign Whole Peak List: Assign all the peaks in the selected peak list to all of their NOE assignment possibilities; only useful if strict tolerances are set