We presented here the KiSSim (Kinase Structural Similarity) fingerprint as a novel structure-enabled pocket encoding tailored to kinase pockets. The fingerprint encodes physicochemical and spatial properties of the 85 KLIFS residues, which are aligned across the structurally covered kinome. On the one hand, the majority of physicochemical bits -size, HBD, HBA, charge, aromatic, and aliphatic, which are adapted from the SiteAlign method -encode the pocket sequence in a generalized, pharmacophoric way. On the other hand, the side chain orientation, solvent exposure, and the spatial bits -the distances to the pocket center and key subpocket centers and the distance distributions' moments -account for the structural conformation. Across all fingerprints, we saw that the fingerprint captures the physicochemical property variability (e.g., most residues are uncharged, whereas HBD/HBA features vary) and the conserved residue positions (e.g., distances to DFG region are more widely spread than to the hinge region). We used the fingerprint to calculate all-against-all distances -small distances refer to high similarity, large distances to low similarity -within the structurally covered kinome: the DFG-in and DFG-out dataset consist of 4112 and 406 structures, representing 257 and 71 kinases, respectively. We found that the fingerprint can distinguish between intra-and inter-kinase similarities and between DFG-in and DFG-out structures. Some kinases are represented by multiple structures, hence some kinase pairs are represented by multiple structure pairs. The distribution of structure distances for one kinase pair can be broad; we selected per kinase pair the closest structure pair that is experimentally observed. We clustered the resulting kinase distance matrix to produce a KiSSim-based kinome tree. While the tree reproduced large parts of the sequence-based Manning tree, some relationships could be observed that are unexpected from a sequence perspective only. For example, we found similarities between CaMKK2 (STE) and DRAK2 (CAMK), which are targeted by the same chemical probe SGC-STK17B-1; 9 we also could confirm the reassignment of AurA, AurC, PLK4, and CaMMK2 from the Other to the CAMK group as proposed by Modi and Dunbrack 7 .