-\subsection{Disorder prediction results}
-Each service operates on sequences in the alignment to identify regions likely
-to be unstructured or flexible, or alternately, fold to form globular domains.
-As a consequence, disorder predictor results include both sequence features and
-sequence associated alignment annotation rows. Section \ref{featannot} describes
-the manipulation and display of these data in detail, and {\bf Figure
-\ref{alignmentdisorder}} demonstrates how sequence feature shading and
-thresholding (described in Section \ref{featureschemes}) can be used to
-highlight differences in disorder prediction across aligned sequences.
-
-\begin{figure}[htbp]
-\begin{center}
-\includegraphics[width=5in]{images/disorderpred.pdf}
-\caption{{\bf Shading alignment by sequence disorder}. Alignment of Interleukin IV homologs coloured with Blosum62 with protein disorder prediction sequence features overlaid, shaded according to their score. Borderline disordered regions appear white, reliable predictions are either Green or Brown depending on the type of disorder prediction. }
-\label{alignmentdisorder}
-\end{center}
-\end{figure}
-
-\subsubsection{Navigating large sets of disorder predictions}
-
-{\bf Figure \ref{alignmentdisorderannot}} shows a single sequence annotated with
-a range of disorder predictions. Disorder prediction annotation rows are
-associated with a sequence in the same way as secondary structure prediction
-results. When browsing an alignment containing large numbers of disorder
-prediction annotation rows, clicking on the annotation row label will highlight
-the associated sequence in the alignment display, and double clicking will
-select that sequence.
-
-\begin{figure}[htbp]
-\begin{center}
-\includegraphics[width=5in]{images/disorderpredannot.pdf}
-\caption{{\bf Annotation rows for several disorder predictions on a sequence}. A
-zoomed out view of a prediction for a single sequence. The sequence is shaded to highlight disordered regions (brown and grey), and the line plots below the Sequence show the raw scores for various disorder predictors. Horizontal lines on each graph mark the level at which disorder predictions become significant. }
-\label{alignmentdisorderannot}
-\end{center}
-\end{figure}
-
-
-\subsection{Disorder predictors provided by JABAWS 2.0}
-For full details of each predictor and the results that Jalview can display,
-please consult
-\href{http://www.jalview.org/help/html/webServices/proteinDisorder.html}{Jalview's
-protein disorder service documentation}. Short descriptions of the methods provided in JABAWS 2.0 are given below:
-
-\subsubsection{DisEMBL}
-\href{http://dis.embl.de/}{DisEMBL (Linding et al., 2003)} is a set of machine-learning based predictors trained to
-recognise disorder-related annotation found on PDB structures.
-
-\textbf{COILS} Predicts
-loops/coils according to DSSP
-definitions\footnote{DSSP Classifications of secondary structure are: $\alpha$-helix (H), 310-helix (G), $\beta$-strand (E)
-are ordered, and all other states ($\beta$-bridge (B), $\beta$-turn (T), bend (S),
-$\pi$-helix (I), and coil (C)) considered loops or coils.}. Features mark range(s) of
-residues predicted as loops/coils, and annotation row gives raw value
-for each residue. Value over 0.516 indicates loop/coil.
-
-\textbf{HOTLOOPS} constitute a refined subset of \textbf{COILS}, namely those loops with
-a high degree of mobility as determined from C$\alpha$ temperature factors (B
-factors). It follows that highly dynamic loops should be considered
-protein disorder. Features mark range(s) of residues predicted to
-be hot loops and annotation row gives raw value for each
-residue. Values over 0.6 indicates hot loop.
-
-\textbf{REMARK465} ``Missing
-coordinates in X-ray structure as defined by remark465 entries in PDB.
-Nonassigned electron densities most often reflect intrinsic disorder,
-and have been used early on in disorder prediction.'' Features give
-range(s) of residues predicted as disordered, and annotation rows gives
-raw value for each residue. Values over 0.1204 indicates disorder.
-
-\subsubsection{RONN {\sl a.k.a.} Regional Order Neural Network}
-\href{http://www.strubi.ox.ac.uk/RONN}{RONN} employs an approach
-known as the `bio-basis' method to predict regions of disorder in
-sequences based on their local similarity with a gold-standard set of
-disordered protein sequences. It yields a set of disorder prediction
-scores, which are shown as sequence annotation below the alignment.
-
-\textbf{JRonn}\footnote{JRonn denotes the score for this server because JABAWS
-runs a Java port of RONN developed by Peter Troshin and distributed as
-part of \href{http://www.biojava.org/}{Biojava 3}} Annotation Row gives RONN score for each residue in
-the sequence. Scores above 0.5 identify regions of the protein likely
-to be disordered.
-
-\subsubsection{IUPred}
-\href{http://iupred.enzim.hu/Help.php}{IUPred} employs
-an empirical model to estimate likely regions of disorder. There are
-three different prediction types offered, each using different
-parameters optimized for slightly different applications. It provides
-raw scores based on two models for predicting regions of `long
-disorder' and `short disorder'. A third predictor identifies regions
-likely to form structured domains.
-
-\textbf{Long disorder} Annotation rows predict
-context-independent global disorder that encompasses at least 30
-consecutive residues of predicted disorder. A 100 residue
-window is used for calculation. Values above 0.5 indicates the residue is
-intrinsically disordered.
-
-\textbf{Short disorder} Annotation rows predict for short, (and
-probably) context-dependent, disordered regions, such as missing
-residues in the X-ray structure of an otherwise globular protein.
-Employs a 25 residue window for calculation, and includes adjustment
-parameter for chain termini which favors disorder prediction at the
-ends. Values above 0.5 indicate short-range disorder.
-
-\textbf{Structured domains} are marked with sequence Features. These highlight
-likely globular domains useful for structure genomics investigation. Post-analysis of disordered region profile to find continuous regions
-confidently predicted to be ordered. Neighbouring regions close to
-each other are merged, while regions shorter than the minimal domain
-size of at least 30 residues are ignored.
-
-\subsubsection{GLOBPLOT}
-\href{http://globplot.embl.de/}{GLOBPLOT} defines regions of
-globularity or natively unstructured regions based on a running sum of
-the propensity of residues to be structured or unstructured. The
-propensity is calculated based on the probability of each amino acid
-being observed within well defined regions of secondary structure or
-within regions of random coil. The initial signal is smoothed with a
-Savitzky-Golay filter, and its first order derivative
-computed. Residues for which the first order derivative is positive
-are designated as natively unstructured, whereas those with negative
-values are structured.
-
-{\bf Disordered region} sequence features are created marking mark range(s) of residues with positive first order derivatives, and
-\textbf{Globular Domain} features mark long stretches of order. \textbf{Dydx} annotation rows gives the first order derivative of smoothed score. Values above 0 indicates
-residue is disordered.
-
-\textbf{Smoothed Score and Raw Score} annotation rows give the smoothed and raw scores used to create the differential signal that
-indicates the presence of unstructured regions. These are hidden
-by default, but can be shown by right-clicking on the alignment
-annotation panel and selecting \textbf{Show hidden annotation}.
-
-\exercise{Protein Disorder Prediction}{
-\label{protdispredex}
-
-\exstep{Open the alignment at
-\url{http://www.jalview.org/tutorial/interleukin7.fa}. }
-
-\exstep{Run the DISEMBL disorder predictor {\slvia} the {\slWeb Services
-$\Rightarrow$ Disorder Prediction } submenu.}
-
-\exstep{Use {\sl Sequence ID $\Rightarrow$ Structure $\Rightarrow$ Discover PDB
-IDs} to retrieve all the PDB structures for the sequences.}
-
-\exstep{Open and align
-the structures for all sequences.
-
-{\sl Hint: see \ref{viewAllStructures} to see how to do this.}}
-
-\exstep{Compare the disorder predictions to the structure data by mapping any
-available temperature factors to the alignment {\sl via} the {\sl Sequence ID
-Popup $\Rightarrow$ Selection $\Rightarrow$ Add reference annotation} option.}
-
-\exstep{Apply the IUPred disorder prediction method}
-\exstep{Use the {\sl Per
-sequence option} in the {\sl Colour $\Rightarrow$ By annotation \ldots} dialog to shade
-the sequences by the long and short disorder predictors.
-Do the two methods agree with the structure ?}}
-
-\section{Features and Annotation}
-\label{featannot}
-Features and annotations are additional information that is overlaid on the sequences and the alignment. Generally speaking, annotations are associated with columns in the alignment. Features are associated with specific residues in the sequence.
-
-Annotations are shown below the alignment in the annotation panel, and often reflect properties of the alignment as a whole. The Conservation, Consensus and Quality scores are examples of dynamic annotation, so as the alignment changes, they change along with it. Conversely, sequence features are properties of the individual sequences, so they do not change with the alignment, but are shown mapped on to specific residues within the alignment.
-
-Features and annotation can be interactively created, or retrieved from external
-data sources. DAS (the Distributed Annotation System) is the primary source of
-sequence features, whilst webservices like JNet (see \ref{jpred} above) can be used to analyse a given sequence or alignment and generate annotation for it.
-
-
-\subsection{Creating sequence features}
-Sequence features can be created simply by selecting the area in a sequence (or sequences) to form the feature and selecting {\sl Selection $\Rightarrow$ Create Sequence Feature } from the right-click context menu (Figure \ref{features}). A dialogue box allows the user to customise the feature with respect to name, group, and colour. The feature is then associated with the sequence. Moving the mouse over a residue associated with a feature brings up a tool tip listing all features associated with the residue.
-
-\begin{figure}[htbp]
-\begin{center}
-\includegraphics[width=2in]{images/feature1.pdf}
-\includegraphics[width=2.5in]{images/feature2.pdf}
-\includegraphics[width=1.5in]{images/feature3.pdf}
-\caption{{\bf Creating sequence features.} Features can readily be created from selections via the context menu and are then displayed on the sequence. }
-\label{features}
-\end{center}
-\end{figure}
-
-Creation of features from a selection spanning multiple sequences results in the creation of one feature per sequence. Each feature remains associated with it's own sequence.
-
-\subsection{Customising feature display}
-
-Feature display can be toggled on or off by selecting the {\sl View
-$\Rightarrow$ Show Sequence Features} menu option. When multiple features are
-present it is usually necessary to customise the display. Jalview allows the
-display, colour, rendering order and transparency of features to be modified
-{\sl via} the {\sl View $\Rightarrow$ Feature Settings\ldots} menu option. This
-brings up a dialogue window (Figure \ref{custfeat}) which allows the
-visibility of individual feature types to be selected, colours changed (by
-clicking on the colour of each sequence feature type) and the rendering order
-modified by dragging feature types to a new position in the list. Dragging the
-slider alters the transparency of the feature rendering. The Feature
-Settings dialog also includes functions for more advanced feature shading
-schemes and buttons for sorting the alignment according to the distribution of
-features. These capabilities are described further in sections
-\ref{featureschemes} and \ref{featureordering}.
-
-\begin{figure}[htbp]
-\begin{center}
-\includegraphics[width=4in]{images/features4.pdf}
-\caption{{\bf Multiple sequence features.} An alignment with JPred secondary structure prediction annotation below it, and many sequence features overlaid onto the aligned sequences. The tooltip lists the features annotating the residue below the mouse-pointer.}
-\end{center}
-\end{figure}
-
-\begin{figure}[htbp]
-\begin{center}
-\includegraphics[width=4in]{images/features5.pdf}
-\caption{{\bf Customising sequence features.} Features can be recoloured, switched on or off and have the rendering order changed. }
-\label{custfeat}
-\end{center}
-\end{figure}
-
-\subsection{Sequence Feature File Formats}
-
-Jalview supports the widely used GFF tab delimited format\footnote{see
-http://www.sanger.ac.uk/resources/software/gff/spec.html} and its own Jalview
-Features file format for the import of sequence annotation. Features and
-alignment annotation are also extracted from other formats such as Stockholm,
-and AMSA. URL links may also be attached to features. See the online
-documentation for more details of the additional capabilities of the jalview
-features file.
-
-\exercise{Creating features}{
-\exstep{Open the alignment at \textsf{http://www.jalview.org/tutorial/alignment.fa}. We know that the Cysteine residues at columns 97, 102, 105 and 135 are involved in iron binding so we will create them as features. Navigate to column 97, sequence 1. Select the entire column by clicking in the ruler bar. Then right-click on the selection to bring up the context menu and select {\sl Selection $\Rightarrow$ Create Sequence Feature}. A dialogue box will appear.
-}
-\exstep{
-Enter a suitable Sequence Feature Name (e.g. ``Iron binding site") in the
-appropriate box. Click on the Feature Colour bar to change the colour if
-desired, add a short description (``One of four Iron binding Cysteines") and press OK. The features will then appear on the sequences. } \exstep{Roll the mouse cursor over the new features. Note that the position given in the tool tip is the residue number, not the column number. To demonstrate that there is one feature per sequence, clear all selections by pressing [ESC] then insert a gap in sequence 3 at column 95. Roll the mouse over the features and you will see that the feature has moved with the sequence. Delete the gap you created.
-}
-\exstep{
-Add a similar feature to column 102. When the feature dialogue box appears, clicking the Sequence Feature Name box brings up a list of previously described features. Using the same Sequence Feature Name allows the features to be grouped.}
-\exstep{Select {\sl View $\Rightarrow$ Feature Settings\ldots} from the
-alignment window menu. The Sequence Feature Settings window will appear. Move
-this so that you can see the features you have just created. Click the check
-box for ``Iron binding site" under {\sl Display} and note that display of this
-feature type is now turned off. Click it again and note that the features are
-now displayed. Close the sequence feature settings box by clicking OK or
-Cancel.} }
-
-\subsection{Creating user defined annotation}
-
-Annotations are properties that apply to the alignment as a whole and are visualized on rows in the annotation panel.
-To create a new annotation row, right click on the annotation label panel and select the {\sl Add New Row} menu option (Figure \ref{newannotrow}). A dialogue box appears. Enter the label to use for this row and a new row will appear.
-
-\begin{figure}[htbp]
-\begin{center}
-\includegraphics[width=1.3in]{images/annots1.pdf}
-\includegraphics[width=2in]{images/annots2.pdf}
-\caption{{\bf Creating a new annotation row.} Annotation rows can be reordered by dragging them to the desired place.}
-\label{newannotrow}
-\end{center}
-\end{figure}
-
-To create a new annotation, first select all the positions to be annotated on the appropriate row. Right-clicking on this selection brings up the context menu which allows the insertion of graphics for secondary structure ({\sl Helix} or {\sl Sheet}), text {\sl Label} and the colour in which to present the annotation (Figure \ref{newannot}). On selecting {\sl Label} a dialogue box will appear, requesting the text to place at that position. After the text is entered, the selection can be removed and the annotation becomes clearly visible\footnote{When annotating a block of positions, the text can be partly obscured by the selection highlight. Pressing the [ESC] key clears the selection and the label is then visible.}. Annotations can be coloured or deleted as desired.
-
-\begin{figure}[htbp]
-\begin{center}
-\includegraphics[width=2in]{images/annots3.pdf}
-\includegraphics[width=2in]{images/annots4.pdf}
-\includegraphics[width=2in]{images/annots5.pdf}
-\caption{{\bf Creating a new annotation.} Annotations are created from a selection on the annotation row and can be coloured as desired.}
-\label{newannot}
-\end{center}
-\end{figure}