Titin and twitchin are giant proteins expressed in muscle. They are mainly composed of domains belonging to the fibronectin class III and immunoglobulin c2 families, repeated many times. In addition, both proteins have a protein kinase domain near the C-terminus. This paper explores the evolution of these and related muscle proteins in an attempt to determine the order of events that gave rise to the different repeat patterns and the order of appearance of the proteins. Despite their great similarity at the level of sequence organization, titin and twitchin diverged from each other at least as early as the divergence between vertebrates and nematodes. Most of the repeating units in titin and twitchin were estimated to derive from three original domains. Chicken smooth-muscle myosin light-chain kinase (smMLCK) also has a kinase domain, several immunoglobulin domains, and a fibronectin domain. From a comparison of the kinase domains, titin is predicted to have appeared first during the evolution of the family, followed by twitchin and with the vertebrate MLCKs last to appear. The so-called C-protein from chicken is also a member of this family but has no kinase domain. Its origin remains unclear but it most probably pre-dates the titin/twitchin duplication.