The cis-defensins are a superfamily of small, cationic, cysteine-rich proteins, sharing a common scaffold, but highly divergent sequences and varied functions from host-defence to signalling. Superfamily members are most abundant in plants (with some genomes containing hundreds of members), but are also found across fungi and invertebrates. However, of the thousands of cis-defensin sequences in databases, only have a handful have solved structures or assigned activities. Non-phylogenetic sequence-analysis methods are therefore necessary to use the relationships within the superfamily to classify members, and to predict and engineer functions.