The structure and assembly dynamics of plasmid actin AlfA imply a novel mechanism of DNA segregation.
Bacterial cytoskeletal proteins participate in a variety of processes, including cell division and DNA segregation. Polymerization of one plasmid-encoded, actin-like protein, ParM, segregates DNA by pushing two plasmids in opposite directions and forms the current paradigm for understanding active plasmid segregation. An essential feature of ParM assembly is its dynamically instability, the stochastic switching between growth and disassembly. It is unclear whether dynamic instability is an essential feature of all actin-like protein-based segregation mechanisms or whether bacterial filaments can segregate plasmids by different mechanisms. We expressed and purified AlfA, a plasmid-segregating actin-like protein from Bacillus subtilis, and found that it forms filaments with a unique structure and biochemistry; AlfA nucleates rapidly, polymerizes in the presence of ATP or GTP, and forms highly twisted, ribbon-like, helical filaments with a left-handed pitch and protomer nucleotide binding pockets rotated away from the filament axis. Intriguingly, AlfA filaments spontaneously associate to form uniformly sized, mixed-polarity bundles. Most surprisingly, our biochemical characterization revealed that AlfA does not display dynamic instability and is relatively stable in the presence of diphosphate nucleotides. These results (i) show that there is remarkable structural diversity among bacterial actin filaments and (ii) indicate that AlfA filaments partition DNA by a novel mechanism.
Polka
Jessica K
JK
Cellular and Molecular Pharmacology, University of California, San Francisco, CA 94158, USA.
Kollman
Justin M
JM
Agard
David A
DA
Mullins
R Dyche
RD
D000199:Actins / Q000737:chemistry* / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000378:metabolism; D001412:Bacillus subtilis / Q000235:genetics / Q000378:metabolism*; D001426:Bacterial Proteins / Q000737:chemistry / Q000378:metabolism; D002455:Cell Division; D004269:DNA, Bacterial / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial / Q000502:physiology; D006160:Guanosine Triphosphate / Q000378:metabolism; D015394:Molecular Structure; D010957:Plasmids / Q000378:metabolism*; D011487:Protein Conformation; D021122:Protein Subunits
D000199:Actins; D001426:Bacterial Proteins; D004269:DNA, Bacterial; D021122:Protein Subunits; D006160:Guanosine Triphosphate; D000255:Adenosine Triphosphate