| Abstract:ABSTRACT: BACKGROUND: Capping protein (CP), also known as CapZ in muscle cells and Cap32/34 in Dictyostelium discoideum, plays a major role in regulating actin filament dynamics. CP is a ubiquitously expressed heterodimer comprising an alpha- and beta-subunit. It tightly binds to the fast growing end of actin filaments, thereby functioning as a "cap" by blocking the addition and loss of actin subunits. Vertebrates contain two somatic variants of CP, one being primarily found at the cell periphery of non-muscle tissues while the other is mainly localized at the Z-discs of skeletal muscles. RESULTS: To elucidate structural and functional differences between cytoplasmic and sarcomercic CP variants, we have solved the atomic structure of Cap32/34 (32 = beta- and 34 = alpha-subunit) from the cellular slime mold Dictyostelium at 2.2 A resolution and compared it to that of chicken muscle CapZ. The two homologs display a similar overall arrangement including the attached alpha-subunit C-terminus (alpha-tentacle) and the flexible beta-tentacle. Nevertheless, the structures exhibit marked differences suggesting considerable structural flexibility within the alpha-subunit. In the alpha-subunit we observed a bending motion of the beta-sheet region located opposite to the position of the C-terminal beta-tentacle towards the antiparallel helices that interconnect the heterodimer. Recently, a two domain twisting attributed mainly to the beta-subunit has been reported. At the hinge of these two domains Cap32/34 contains an elongated and highly flexible loop, which has been reported to be important for the interaction of cytoplasmic CP with actin and might contribute to the more dynamic actin-binding of cytoplasmic compared to sarcomeric CP (CapZ). CONCLUSIONS: The structure of Cap32/34 from Dictyostelium discoideum allowed a detailed analysis and comparison between the cytoplasmic and sarcomeric variants of CP. Significant structural flexibility could particularly be found within the alpha-subunit, a loop region in the beta-subunit, and the surface of the alpha-globule where the amino acid differences between the cytoplasmic and sarcomeric mammalian CP are located. Hence, the crystal structure of Cap32/34 raises the possibility of different binding behaviours of the CP variants toward the barbed end of actin filaments, a feature, which might have arisen from adaptation to different environments.
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