Engineering indel and substitution variants of diverse and ancient enzymes using Graphical Representation of Ancestral Sequence Predictions (GRASP)
Fig 6
A tree with five extant sequences and ancestors with bi-directional and uni-directional support for different edges shown.
A, Input POG and the matrix E* (recovering all edges under consideration for ancestors). Red squares refer to all edges involving position 5 (occupied by W in all sequences). Non-zero entries in the red squares identify indices for 2 and 4, as well as the end terminus. B, Ancestor POGs (labelled N0-N3) are shown at branch points as inferred by bi-directional edge parsimony. Solid arrows indicate bi-directional support; dashed arrows indicate uni-directional support. For uni-directional edges, the direction of support is shown by the direction of the arrow. The N1 ancestor is annotated to illustrate the relationship between indices in an ancestral matrix and the edges in the resulting ancestor POGs. The (a, b) and (b, a) indices are highlighted for select positions—for example, (4,5) and (5,4) are the forwards support from index 4—5 and backwards support from index 5—4 (green boxes and green edge in N1) showing that the edge from 4—5 is supported in both directions. The edge from 2—5 is supported going forward but not backward (blue boxes and blue edge in N1). Other edges forward from index 2 are maximally parsimonious in the forward direction at N1, such as 2—3 and 2—4, but only the support for 4 is reciprocated (purple boxes and purple edges in N1) recovering a “preferred” path from 2 to 4 to 5 in N1. C, Extant POGs, with position indices.