Assignment & Structure Validation

Introduction

The final part of these tutorial exercises is to look at how we can use the CING software to analyse and validate NMR and structure data. Also, we will consider the data checks that can be performed within Analysis, without the need for external programs, for example to find chemical shift outliers and bogus assignments. This part of the Tutorial is based on the CcpnStructCourseD project.

Structure Validation

In the main menu select  "M: Structure: CING: Validate Structures" and click the green [New Run] button in the upper right corner. This run specification will contain all of the data that CING will analyse. When a new run is made the {Structures} tab will fill with the various models from an ensemble. Accepting this, we move on to the {Shifts & Measurements} tab. Here click [Add Measurement List] to add the shift list to the CING analysis. Move to the {Peak Lists} tab, and add the N-NOESY:182:2 peak list (this is the one that ARIA made) by selecting it from the lower right pulldown menu and selecting [Add Peak List]. Finally in the restraint lists tab add restraint list "Distance-2:1". This restraint list is the one that came back from ARIA and consequently lives in the second restraint set. Note that in general you may select as much data as you like, but we are just selecting a smaller subset for demonstration purposes.

Select the {Run Settings} tab. To submit the analysis press the [Submit Project] button. Note that this may take some time, so you may wish to let the demonstrator make a real submission and view some previously calculated results, thus sparing the iCING server. Although, feel free to use this server generally.

Assignment Quality Control

At the menu option M: Assignment: Quality Reports you can access a suite of analyses which generate automated reports about the quality and completeness of your assignments. The first tab simply lists how many of each kind of residue and atom type you have resonance assignments for. The second tab shows statistics for how many NOE (and other through-space) connections you have across your molecular system.

While the first two tabs are dedicated to analysing completeness, the last two tabs are dedicated to finding mistakes and inconsistencies in assignments. These tables look at the same kind of issues, but one presents the information from a resonance point of view, and the other from a peak point of view. Most notably the assignments are checked for the following criteria:

• • An assignment to a given atom is duplicated.

  • The standard deviation for a chemical shift is large.

  • If a peak position is far from the chemical shift average (irrespective of a shift's deviation).

  • A resonance has an impossible number of covalently bound partners: e.g. an amide proton assigned to two HSQC peaks cannot be linked to both nitrogen resonances.

  • The chemical shift for an atom assignment is unlikely given the known chemical shift distributions (BMRB).

  • If peaks have unusual intensities or sign.

When something is potentially wrong, then cells of the tables are coloured red. A highlighted cell doesn't mean that something is definitely wrong, just that it warrants an explanation or further investigation. To find the peaks responsible for any aberration, simply click on the row in the table and click [Show Peaks]. In the resulting table you can assign or find any of the peaks in the various spectrum windows.

Previous: Using Structures to Assign NOE Contributions

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