Leiomodins are larger homologs of tropomodulin, a tropomyosin-binding, actin-capping protein. There are several leiomodin isoforms, one of them found in smooth muscles (Lmod1) and another one found in cardiac and skeletal muscles (Lmod2). In this work, the tropomyosin-binding abilities of these two isoforms were studied. The tropomyosin-binding sites were localized in the N-terminal regions of Lmod1 and Lmod2. The affinities of the leiomodin fragments containing the tropomyosin-binding sites for tropomyosin peptides containing N-termini of different tropomyosin isoforms, α, γ and δ, were determined and compared using non-denaturing gel-electrophoresis and circular dichroism. It was shown that leiomodin/tropomyosin binding is isoform-specific and differs almost 100-fold for different tropomyosin isoforms.