Characterization of DNA/RNA binding properties of CspA protein from Mycobacterium tuberculosis

• Complexes of MtbCspA with ssDNA are more stable than with ssRNA. • MtbCspA does not appear to bind RNA hairpins. • The DNA/RNA binding site of MtbCspA includes β3-β4 and β4-β5 loops. • Our data show that MtbCspA apparently cannot function as an RNA chaperone. CspA from Mycobacterium tuberculosis (MtbCspA) is a classical small DNA/RNA-binding protein that has the potential to interact with non-coding sRNA, thereby participating in bacterial host adaptation and pathogenicity. We used molecular dynamics simulations coupled with EMSA to test the ability of MtbCspA to bind single-stranded DNA and RNA, as well as short hairpin RNA. We found that MtbCspA has an extended and relatively large DNA/RNA binding region compared to previously defined ones. This area includes classical RNP1/RNP2 motifs and protruding β3-β4 and β4-β5 loops. The protein has a lower affinity for ssRNA compared to ssDNA and poor affinity for the tested RNA hairpins. These results suggest that CspA may not be a fully functional RNA chaperone in M. tuberculosis but may be more involved in transcriptional regulation in this bacterium.