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Formation of stable actin filaments, critically important for actin functions, is determined by the ionic strength of the solution. However, not much is known about the elements of the actin fold involved in ionic-strength-dependent filament stabilization. In this work, F-actin was destabilized by Cu2+ binding to Cys374, and the effects of solvent conditions on the dynamic properties of F-actin were correlated with the involvement of Segment 227-235 in filament stabilization. The results of our work show that the presence of Mg2+ at the high-affinity cation binding site of Cu-modified actin polymerized with MgCl2 strongly enhances the rate of filament subunit exchange and promotes the filament instability. In the presence of 0.1 M KCl, the filament subunit exchange was 2–3-fold lower than that in the MgCl2-polymerized F-actin. This effect correlates with the reduced accessibility of the D-loop and Segment 227-235 on opposite filament strands, consistent with an ionic-strength-dependent conformational change that modulates involvement of Segment 227-235 in stabilization of the intermonomer interface. KCl may restrict the mobility of the α-helix encompassing part of Segment 227-235 and/or be bound to Asp236 at the boundary of Segment 227-235. These results provide experimental evidence for the involvement of Segment 227-235 in salt-induced stabilization of contacts within the actin filament and suggest that they can be weakened by mutations characteristic of actin-associated myopathies.

The article reviews the discovery, properties and functional activities of new bacterial enzymes, proteases grimelysin (ECP 32) of Serratia grimesii and protealysin of Serratia proteamaculans, characterized by both a highly specific “actinase” activity and their ability to stimulate bacterial invasion. Grimelysin cleaves the only polypeptide bond Gly42-Val43 in actin. This bond is not cleaved by any other proteases and leads to a reversible loss of actin polymerization. Similar properties were characteristic for another bacterial protease, protealysin. These properties made grimelysin and protealysin a unique tool to study the functional properties of actin. Furthermore, bacteria Serratia grimesii and Serratia proteamaculans, producing grimelysin and protealysin, invade eukaryotic cells, and the recombinant Escherichia coli expressing the grimelysin or protealysins gene become invasive. Participation of the cellular c-Src and RhoA/ROCK signaling pathways in the invasion of eukaryotic cells by S. grimesii was shown, and involvement of E-cadherin in the invasion has been suggested. Moreover, membrane vesicles produced by S. grimesii were found to contain grimelysin, penetrate into eukaryotic cells and increase the invasion of bacteria into eukaryotic cells. These data indicate that the protease is a virulence factor, and actin can be a target for the protease upon its translocation into the host cell.

Formation of stable actin filaments, critically important for actin functions, is determined by the ionic strength of the solution. However, not much is known about the elements of the actin fold involved in ionic-strength-dependent filament stabilization. In this work, F-actin was destabilized by Cu2+ binding to Cys374, and the effects of solvent conditions on the dynamic properties of F-actin were correlated with the involvement of Segment 227-235 in filament stabilization. The results of our work show that the presence of Mg2+ at the high-affinity cation binding site of Cu-modified actin polymerized with MgCl2 strongly enhances the rate of filament subunit exchange and promotes the filament instability. In the presence of 0.1 M KCl, the filament subunit exchange was 2–3-fold lower than that in the MgCl2-polymerized F-actin. This effect correlates with the reduced accessibility of the D-loop and Segment 227-235 on opposite filament strands, consistent with an ionic-strength-dependent conformational change that modulates involvement of Segment 227-235 in stabilization of the intermonomer interface. KCl may restrict the mobility of the α-helix encompassing part of Segment 227-235 and/or be bound to Asp236 at the boundary of Segment 227-235. These results provide experimental evidence for the involvement of Segment 227-235 in salt-induced stabilization of contacts within the actin filament and suggest that they can be weakened by mutations characteristic of actin-associated myopathies. [ABSTRACT FROM AUTHOR]

The article reviews the discovery, properties and functional activities of new bacterial enzymes, proteases grimelysin (ECP 32) of Serratia grimesii and protealysin of Serratia proteamaculans, characterized by both a highly specific "actinase" activity and their ability to stimulate bacterial invasion. Grimelysin cleaves the only polypeptide bond Gly42-Val43 in actin. This bond is not cleaved by any other proteases and leads to a reversible loss of actin polymerization. Similar properties were characteristic for another bacterial protease, protealysin. These properties made grimelysin and protealysin a unique tool to study the functional properties of actin. Furthermore, bacteria Serratiagrimesii and Serratia proteamaculans, producing grimelysin and protealysin, invade eukaryotic cells, and the recombinant Escherichia coli expressing the grimelysin or protealysins gene become invasive. Participation of the cellular c-Src and RhoA/ROCK signaling pathways in the invasion of eukaryotic cells by S. grimesii was shown, and involvement of E-cadherin in the invasion has been suggested. Moreover, membrane vesicles produced by S. grimesii were found to contain grimelysin, penetrate into eukaryotic cells and increase the invasion of bacteria into eukaryotic cells. These data indicate that the protease is a virulence factor, and actin can be a target for the protease upon its translocation into the host cell. [ABSTRACT FROM AUTHOR]