Inhibitory effect of melittin on endonuclease-like activity of centrin

Abstract

The xeroderma pigmentosum group C protein (XPC) and centrin2 are the primary initiators of global genome nucleotide excision repair (NER). Centrin, acts as a member of the EF-hand super family of calcium-binding proteins, playing roles in reconstitution of the vitro NER reaction. To understand the possible molecular and structural properties of the multiprotein process, the interactions of Euplotes octocarinatus centrin (EoCen), melittin, and DNA are described. EoCen shares a sequence identity of 66% with centrin2. Melittin possesses inverse direction hydrophobic triads-leucine-leucine-tryptophan (LLW) which are responsible for centrin binding. It is applied as a natural peptide to mimic centrin target peptide. As a result, it is proved that the integrated protein shows an endonuclease-like activity to DNA. Melittin is capable of interaction with both EoCen and DNA. More importantly, it is found that melittin displays an inhibitory effect on the endonuclease-like activity of centrin when it co-exists with EoCen and DNA in solution. Meanwhile, the DNA-melittin-EoCen ternary complex forms in the process. Quantitative analyses demonstrated by extensive biophysical assays reveal that binding of the peptide to DNA or centrin modulates the binding properties of it to another component. Furthermore, a possible positioning model of DNA and EoCen on melittin is proposed. This finding may constitute a model for that existing between centrin and its target peptide in NER process.