Effect of divalent ions on the minimal relaxase domain of MobA.
The MobA protein encoded by plasmid R1162 plays an important role in conjugative mobilization between bacterial cells. It has two functional domains, the N-terminal relaxase domain and C-terminal primase domain. The N-terminal 186 residues (minMobA) is the minimal domain required for relaxase activity. We investigated the effects of different divalent metallic cations on minMobA activity measuring DNA binding, DNA nicking, and protein denaturation experiments. The results show that divalent cations are not required for DNA binding but are required for DNA nicking. The range of metals that function in minMobA suggests the cation role is largely structural. The most tightly binding cation is Mn(2+), but the expressed protein shows roughly equal amounts of Mg(2+) and Ca(2+), both of which facilitate substrate binding and catalysis. Surprisingly, Zn(2+) does not facilitate DNA binding nor allow nicking activity.
Xia
Shuangluo
S
Institute for Cellular and Molecular Biology, Department of Chemistry and Biochemistry, University of Texas, Austin, TX 78712, USA.
Robertus
Jon D
JD
D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D001483:Base Sequence; D055162:Biocatalysis / Q000187:drug effects; D002151:Calorimetry; D020134:Catalytic Domain / Q000187:drug effects; D002413:Cations, Divalent / Q000494:pharmacology*; D002942:Circular Dichroism; D004277:DNA, Single-Stranded / Q000378:metabolism; D013058:Mass Spectrometry; D017434:Protein Structure, Tertiary; D011550:Pseudomonas aeruginosa
D001426:Bacterial Proteins; D002413:Cations, Divalent; D004277:DNA, Single-Stranded