A biochemically active MCM-like helicase in Bacillus cereus

Abstract

The mini-chromosome maintenance (MCM) proteins serve as the replicative helicases in archaea and eukaryotes. Interestingly, an MCM homolog was identified, by BLAST analysis, within a phage integrated in the bacterium Bacillus cereus (Bc). BcMCM is only related to the AAA region of MCM-helicases; the typical amino-terminus is missing and is replaced by a segment with weak homology to primases. We show that BcMCM displays 3′→5′ helicase and ssDNA-stimulated ATPase activity, properties that arise from its conserved AAA domain. Isolated BcMCM is a monomer in solution but likely forms the functional oligomer in vivo. We found that the BcMCM amino-terminus can bind ssDNA and harbors a zinc atom, both hallmarks of the typical MCM amino-terminus. No BcMCM-catalyzed primase activity could be detected. We propose that the divergent amino-terminus of BcMCM is a paralog of the corresponding region of MCM-helicases. A divergent amino terminus makes BcMCM a useful model for typical MCM-helicases since it accomplishes the same function using an apparently unrelated structure.

Molecular architecture of a multifunctional MCM complex

“…The C-terminal region BcMCM 501–1028 , which contains the helicase activity, is sufficient to bind DNA (Figure 2C). Interestingly, the amino terminal BcMCM 1–361 , where the canonical primase domain is located, did not associate with DNA (Figure 2D) whereas the BcMCM 1–400 construct binds DNA with low affinity, supporting previous results (31). Therefore these experiments suggest that the fragment comprising residues 361–400 contributes to DNA binding by the primase domain.…”

“…A previous report showed that BcMCM binds ssDNA (31). However, BcMCM can also bind DNA probes that contain regions of single and double strand, such as template–primer molecules with 3′ and 5′ overhangs, and bubble-like structures (Figure 2A).…”

Staphylococcal SCCmec elements encode an active MCM-like helicase and thus may be replicative

“…Cch and MCM also both translocate in the 3’ to 5’ direction and both have a C-terminal WH domain. We could find only one other reported prokaryotic MCM-like helicase, BcMCM from a Bacillus cereus bacteriophage, which was identified by its sequence homology to MCMs 41 . Although Cch, BcMCM and archaeal MCM all have different N-terminal domains, their helicase and WH domains probably share a common ancestor.…”

Novel Families of Archaeo-Eukaryotic Primases Associated with Mobile Genetic Elements of Bacteria and Archaea

“…Notably, MCM helicases are generally considered to be specific for eukaryotes and archaea, where they are principal replicative helicases. In the bacterial realm, MCM-family helicases have been previously described only in bacteriophages [9] , [39] , [40] . Our current analysis suggests that MCM-like helicases are also encoded by bacterial (integrative) plasmids (Supplementary table S1), suggesting that MCM-encoding genes are more broadly distributed among bacterial MGE than previously recognized.…”