Molecule : Reference Isotope Schemes

Create and Edit Per-residue Reference Isotope Schemes

This system allows the user to create schemes that describe particular patterns of atomic isotope labelling in terms of combinations of isotopically labelled forms of residues. Once constructed, these schemes may then be applied to a molecule of known residue sequence to gauge the levels of spin-active isotope incorporation in an NMR experiment. This information is useful in several places withing Analysis, including giving more intelligent assignment options and in the generation of distance restraints by matching peak positions to chemical shifts. Although the schemes may be used directly they are typically used as reference information for configuring the Isotope Labelling system; where isotope labelling patterns are tied to particular molecules and experiments.

Because all of the different isotope labelled versions (isotopomers) of each residue type are described independently, a scheme can be used to estimate the specific amounts of incorporation present at multiple atom sites at the same time. For example, although a residue type may have significant levels of 13C at the CA and CB positions on average, there may be no form of the residue where CA and CB are labelled at the same time, and thus CA-CB correlations would not be observed in NMR.

This popup window is divided into three main tabs, the first describes the overall schemes that are available; that would be applied to a molecule in a given situation. The second tab details the residue isotopomer components within the selected scheme, i.e. which labelled residue forms are present. The last tab displays isotopomer labelling in a graphical, three-dimensional way. If any isotope labelling schemes have been created or edited the user may immediately save these to disk via the [Save Schemes] button to the right of the tabs, although these will naturally be saved when the main CCPN project is.

Reference Schemes

This table lists all of the reference isotope schemes that are available to the project. A number of standard schemes are included by default, as part of the main CCPN installation. However, the user is free to create new schemes, either from a completely blank description or by copying and modifying one of the existing schemes. By selecting on a isttope scheme row in the table the scheme is selected to be active for the whole popup and the user can see the contents of the scheme via the other two tabs.

It should be noted that the user cannot edit the standard schemes provided by CCPN, given that these are stored with the software. Any new or copied schemes that the user creates will be stored inside the current CCPN project. If a new scheme should be made available to multiple projects, its XML file can be copied into the main CCPN installation, if the user has appropriate write access.

Isotopomers

The middle tab allows the user to view, and where appropriate edit, the isotope labelling descriptions for the residues within the current scheme (selected in the pulldown menu at the top). An isotope scheme is constructed by specifying one or more isotopomers for each residue type. Each isotopomer represents a different way of incorporating spin-active atom labels into a given kind of residue. Often there will only be one labelled form of a residue, and hence one isotopomer. However, with some kinds of isotope enrichment, for example using glycerol 13C labelled at the C2 position, connected labelled and unlabelled atom sites can be incorporated in alternative ways, resulting in distinct forms of labelling patterns that are not the result of a pure random mix. Knowing which labels are present at the same time, in the same isotopomer form, can be very important for determining which NMR correlations are possible.

In general use when looking through the default, immutable reference schemes that come with CCPN the user can scroll through the isotopomer versions of each residue in the upper table. By clicking on one of these rows the lower table is filled with details of the amount of each kind of isotope (on average) at each atom site. For the lower “Atom Labels” table only one kind of chemical element is shown at a time, but the user may switch to a different one via the “Chemical Element” pulldown.

Editing Isotopomers

When dealing with copied or new isotope schemes the user is allowed to edit all aspects of the scheme. With a completely new scheme there will be no isotopomer records to start with and it is common practice to fill in a standard set of isotopomers, one for each residue type, made with a base level of isotope incorporation. To set this base level the user can use [Set Default Abundances] to manually specify values, although the default is to use natural abundance levels, which is appropriate in most circumstances. With the base levels set the [Add Default Abundance Set] will automatically fill-in a starting set of isotopomers for the scheme. Extra isotopomers can be added for a specific type of residue via the [Add New:] function and adjacent pulldown menu or by copying existing ones; whichever is easier. Each isotopomer has an editable weight to enable the user to indicate the relative abundance within a given residue type.

Once a new isotopomer specification is created clicking on its row allows the user to specify the isotope labelling pattern in the lower “Atom Labels” table. Here the user selects which kind of chemical element to consider and then double-clicks to edit the “Weighting” columns in the table. The weightings represent the relative abundance of a given nuclear isotope at a given atom site. The weightings could be set as ratios, fractions, or percentages; it is only the relative proportion that is important. For example if a carbon atom site was known to have 5% Carbon-12 and 95% Carbon-13 isotopes then the respective weights could be entered as 1 & 19 or 0.05 & 0.95; whatever is most convenient. For efficient setup of schemes the [Propagate Abundances] function can be used to spread the same levels of incorporation over several atom sites (from the last selected row).

Isotopomer Structure

The last tab is an alternative way of presenting the isotope patterns present within the residues of the current scheme (selected in either of the first two tabs). Here the user selects a residue type in the upper left pulldown menu and then a numbered isotopomer, or an average of all isotopomers, in the right hand pulldown menu. The structural display will show a moveable picture of the residue (in a standard conformation) where unlabelled atom sites are represented with grey spheres, labelled sites with yellow spheres and intermediate incorporation with shades in between.

It should be noted that this kind of 3D display is only possible if there is an idealised structure available for a residue type. This data will be present for all of the regular biopolymer residues, but may be missing for more unusual compounds; although a lack of coordinates does not impact upon the isotopomer setup.

To move and rotate the three-dimensional residue display the following keyboard controls may be used:

  • Rotate: Arrow keys
  • Zoom: Page Up & Page Down keys
  • Translate: Arrow keys + Control key

Or alternatively the following mouse controls:

  • Rotate: Middle button click & drag
  • Zoom: Mouse wheel or middle button click + Shift key & drag up/down
  • Translate: Middle button click & drag + Control key

Also an options menu appears when the right mouse button is clicked.

Main Panel

button Save Schemes: Save all changes to the reference isotope scheme to disk; the saves ALL changes to the CCPN installation for all projects to use

button Clone: Clone popup window

button Help: Show popup help document

button Close: Close popup

Reference Schemes

A table of all of the reference isotope scheme definitions available to the project

Table 1
Code A short textual code that identifies the reference isotope scheme in graphical displays
Name The full name for the isotope scheme (Editable)
Description A detailed description of the isotope scheme including user comments (Editable)
Save Location The name of the CCPN data repository in which the isotope scheme is saved; “refData” is in the CCPn installation

button Copy: Make a new reference isotope scheme definition based on a copy of the scheme currently selected

button Delete: Delete the selected isotope scheme

button New: Make a new, blank isotope scheme

Isotopomers

A list of the residue isotopomers that comprise the selected isotope labelling scheme

pulldown Reference Scheme: Selects which of the available isotope schemes to view/edit

Isotopomers

button Set Default Abundances: Opens a panel that allows you to set the basis/default abundances for C, H & N isotopes; used as the starting point for new isotopomer definitions

button Set Natural Abundance Default: Sets the basis/default abundances for C, H & N isotopes to their natural abundance proportions

pulldown Molecule Type: Selects which type of bio-polymer to define residue isotopomer labelling for

Table 2
Ccp Code The CCPN code that identifies the kind of residue the isotopomer relates to
Variant The number of the particular isotopomer (isotope pattern) within its residue type
Weight The fraction of the total residues, of its kind, that the isotopomer make up (Editable)

button Delete Selected: Delete the selected residue isotopomers from the current isotope scheme

button Copy Selected: Make a new residue isotopomer definition by copying the details of the last selected isotopomer

button Add Default Abundance Set: Add a complete set of isotopomers to the isotope scheme, one for each residue type, based on the states default isotope abundances

button Set Atom Type To Default: For all residue isotopomers in the scheme, set the labelling of one kind of atom (the user is prompted) to its default isotopic incorporation

button Add New: Add a new residue isotopomer definition that uses the default isotopic incorporation

pulldown *None*: Selects which kind of residue isotopomer may be added to the current isotope scheme

Atom Labels

pulldown Chemical Element: Selects which kind of atoms to select from the selected residue isotopomer; to display isotopic incorporation in the below table

check Water Exchangeable Atoms: Sets whether to show atoms considered as being “water exchangeable”; their isotopic labelling will rapidly equilibrate with aqueous solvent

Table 3
Atom Name The name of the atom within its residue, for which to set isotopic abundances, within the selected isotopomer
Weighting 12C The amount of 12C isotope incorporation; can be a ratio, percentage or fraction (the value used is relative to the sum of all weights) (Editable)
Weighting 13C The amount of 13C isotope incorporation; can be a ratio, percentage or fraction (the value used is relative to the sum of all weights) (Editable)
%12C The percentage 12C isotope incorporation, calculated using stated weights
%13C The percentage 13C isotope incorporation, calculated using stated weights

button Reset Selected to Default Abundance: For the selected atom sites, in the current isotopomer, set their isotopic incorporation to the default values

button Propagate Abundances: Spread the isotopic incorporation values from the last selected atom site to all selected atoms sites

Isotopomer Structure

A three-dimensional representation of residues and their isotopomer labelling

pulldown Residue Type: Selects which kind of residue, within the current isotope scheme, to show isotopomer structures for

pulldown Isotopomer: Selects which kind of isotopomer (labelling pattern) to display, from the selected residue type.

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