Fig. 5: BPSL1038 may interact with double-stranded DNA via an arginine clamp feature similar to Eco_Cas2.

a The SSM superposition of BPSL1038 dimer (green and cyan for subunit A and B, respectively) with the E. coli Cas2 protein dimer (Eco_Cas2, light pink) that interacts with DNA substrates in the Cas1-Cas2-protospacer DNA Complex (PDB ID: 5DS5). The overall homodimeric structure of BPSL1038 is similar to the Eco_Cas2 with r.m.s.d of 3.1 Å over 123 aligned Cα atoms. The R16 and R78 arginine clamp of Cas2 interacts with protospacer DNA. The residues R10 and R30 from each subunit of BPSL1038 share similar coordination with the ‘arginine clamp’ that may also interact with the DNA substrate. b The Eco_Cas2-DNA complex (PDB ID: 5DS5) is shown in the same orientation as BPSL1038-DNA model (for better comparison, Eco_Cas2 was coloured in green and cyan same as BPSL1038). The side chain rotamer conformation of BPSL1038 R10 and R30 is modelled (yellow) to interact with each strand of DNA. c A close-up view of the modelled dsDNA minor groove interacting with the proposed BPSL1038 ‘arginine clamp’. The D11 and SST active site of BPSL1038 is located between the ‘arginine clamp’ pair, which may accommodate one or two catalytic Mg²⁺ to cleave the DNA backbone before the arginine clamped nucleotide (circled) located close to the D11 aspartate pair (left). The 2Fo-Fc maps for the active site are contoured at 1.5σ. d Schematic presentation of the proposed DNA cleavage mechanism in which the aspartate pair coordinate with a metal cation (Mg²⁺) to activate a nucleophilic water which coordinated with the cation. The activated hydroxyl group of the water molecule will initiate an attack to the scissile phosphate. The negatively charged trigonal bipyramidal transition state is likely stabilized by the metal ion or the side chain of R30. The Mg²⁺ ion may also coordinate with residue S33.