RUVB2

PF3D7_1106000 (RUVB2) is a nuclear AAA+ ATPase of Plasmodium falciparum that couples ATP hydrolysis to weak intrinsic DNA helicase activity, positioning it as an enzymatic component of the parasite's intraerythrocytic developmental program (PMID:24161468). Recombinant and endogenous RUVB2 hydrolyze ATP and unwind DNA in vitro, but the helicase activity is only weak on its own (PMID:24161468). RUVB2 physically associates with PfRuvB3, and specifically not with PfRuvB1, and PfRuvB3 significantly stimulates RUVB2 helicase activity, with the RUVB2/PfRuvB3 complex preferentially translocating and unwinding DNA in the 5'→3' direction (PMID:24161468). RUVB2 is expressed throughout asexual intraerythrocytic development and is predominantly nuclear in merozoite, ring, and trophozoite stages, but partially relocalizes to the cytoplasm during the schizont stage, coinciding with the appearance of the mitosis-specific partner PfRuvB3 (PMID:24161468). Beyond this in vitro enzymatic and interaction characterization, no further mechanistic role, substrate, or in vivo function of RUVB2 has been characterized in the available corpus.

1. 2013 Medium

   Establishing that RUVB2 is itself an enzyme addressed whether the protein carries catalytic activity or is merely a structural complex member, demonstrating it is an ATP-hydrolyzing DNA helicase.

   Evidence In vitro ATPase and DNA helicase assays on recombinant and endogenous RUVB2

   PMID:24161468

   Open questions at the time

   • helicase activity is weak, leaving the physiological substrate unidentified
   • no structural model of the active site or nucleotide-binding fold
   • activity shown only in vitro, not linked to an in vivo DNA transaction
2. 2013 Medium

   Defining RUVB2's binding partner clarified which RuvB-family members it partners with, showing a selective interaction with PfRuvB3 and not PfRuvB1.

   Evidence Immunopurification of the PfRuvB3 complex plus in vitro interaction assay

   PMID:24161468

   Open questions at the time

   • stoichiometry and higher-order architecture of the RUVB2/PfRuvB3 complex not resolved
   • interaction interface not mapped
   • whether additional proteins join this complex in vivo is unknown
3. 2013 Medium

   Reconstituting RUVB2 with PfRuvB3 answered whether the partner modulates catalysis, showing PfRuvB3 stimulates RUVB2 helicase activity and confers 5'→3' directional translocation.

   Evidence In vitro DNA helicase assay with/without PfRuvB3 and directional translocation assay

   PMID:24161468

   Open questions at the time

   • the DNA substrate acted upon in the cell is not identified
   • mechanism by which PfRuvB3 stimulates activity not defined
   • biological process requiring this unwinding activity unknown
4. 2013 Medium

   Mapping RUVB2 expression and localization across the asexual cycle addressed when and where the enzyme acts, showing predominantly nuclear localization that partially shifts to the cytoplasm at schizont stage in concert with PfRuvB3.

   Evidence In vivo immunofluorescence/localization across intraerythrocytic developmental stages

   PMID:24161468

   Open questions at the time

   • functional consequence of the schizont-stage cytoplasmic relocalization unknown
   • no loss-of-function or essentiality data for RUVB2
   • trigger driving the nuclear-to-cytoplasmic shift not identified

• The in vivo DNA transaction, cellular process, and essentiality of RUVB2 remain unresolved.
• no genetic knockout/knockdown phenotype for RUVB2
• physiological DNA substrate and pathway (e.g. replication, repair, chromatin remodeling) not established
• structural basis of RUVB2/PfRuvB3 complex not determined