The aim of the study we present in this PhD thesis is to gain a deeper insight into structure of Z-disc proteins FATZ 1 and y-filamin.
Z-discs are multi-protein complexes which are the primary conduits of the force generated by striated muscle contraction. The protein composition of Z-disc is not well defined and new proteins are continuously being reported.
FATZ 1 is expressed early during myofibrilogenesis and it is presumed to have an important role in Z-disc assembly and functioning. At the moment, it is the only Z-disc protein for which a direct connection with the signal transducer protein phosphatase calcineurin has been demonstrated. The protein sequence of FATZl does not account for its interaction with numerous proteins as it does not contain any known protein interaction domains. For this reason we investigated the structural characteristics of this Z-disc protein.
We studied also y-filamin, a binding partner of FATZl. The role of filamins in cytoskeleton organization and signal transduction has been well documented. y-filamin is the muscle specific isoform of filamins and it has not been studied as extensively as other filamin isoforms. y-filamin can be found both at sarcolemma and at the Z-disc of striated muscle cells and for this reason is considered to be a structural and functional link between contractile apparatus and sarcolemma. y-filamin repeat 23 was reported to be necessary for the interaction of y-filamin and FATZ. We determined crystal structure of y-filamin repeat 23, and found that y-filamin repeat 23 bound nickel.
Detailed analysis of y-filamin protein sequence indicated that other filamin repeats could bind nickel or similar divalent cations. Physiological importance of nickel in Eukaryotes is not well studied, but in Prokaryotes nickel is an essential component of many enzymes. Nickel binding to a protein with mainly structural role would be an exception, since nickel is bound mainly to proteins with enzymatic activities.
We analyzed also structural characteristics of y-filamin repeat 20. The y-filamin repeat 20 has the unique muscle specific insertion responsible for protein localization in the Z-disc. The muscle specific insertion presents no homology with any sequence with known protein structure. Our results from structural studies of repeat 20 indicate that the insertion in repeat 20 influences the fold of this repeat and makes it structurally different from y-filamin repeat 23.