Crystallographic Structures of Titin Immunoglobulin-Like I21 Domains Involved in Dilated Cardiomyopathy

Abstract

The protein titin is the main responsible for the passive elasticity of the sarcomere, determining the stiffness and contractile function of cardiomyocytes. Titin is also a key factor in the etiology of heart disease, given that truncating mutations in the titin gene are the most common cause of dilated cardiomyopathy (DCM). However, missense mutations have commonly been classified as variants of uncertain significance because of their high frequency in the general population and the absence of functional annotation. We have recently identified a missense mutation targeting a conserved cysteine in the I21 domain of titin as a cause of DCM in humans. Here, we present the structural characterization of the wild type domain, that follows a fold characteristic for the intermediate set (type I) of the immunoglobulin superfamily: 2 antiparallel 4-strand-β-sheets packed in a β-sandwich shape. Interestingly, we observed that the I21 titin domain contains a second cysteine located 5Å away from the cysteine position mutated in dilated cardiomyopathy patients. We also report the structural and biophysical characterization of the mutated domain. The mutated domain presents unaltered overall structure, although its thermal stability is severely compromised, showing a decrease of 17ºC in its melting temperature when compared to the wild type domain. Our results suggest that titin domain destabilization by missense variants can lead to DCM, even if the structure of the native state is preserved. Future research will investigate the origin of the thermal destabilization in the mutated domain, and the potential influence of intramolecular disulfide bonds.