Abstract
There have been very few reports on protein domains that specifically recognize sulfur. Here we present the crystal structure of the sulfur-binding domain (SBD) from the DNA phosphorothioation (PT)-dependent restriction endonuclease ScoMcrA. SBD contains a hydrophobic surface cavity that is formed by the aromatic ring of Y164, the pyrolidine ring of P165, and the non-polar side chains of four other residues that serve as lid, base, and wall of the cavity. The SBD and PT-DNA undergo conformational changes upon binding. The S187RGRR191 loop inserts into the DNA major groove to make contacts with the bases of the GPSGCC core sequence. Mutating key residues of SBD impairs PT-DNA association. More than 1000 sequenced microbial species from fourteen phyla contain SBD homologs. We show that three of these homologs bind PT-DNA in vitro and restrict PT-DNA gene transfer in vivo. These results show that SBD-like PT-DNA readers exist widely in prokaryotes.
Highlights
There have been very few reports on protein domains that recognize sulfur
In order to elucidate the molecular mechanism of ScoMcrA recognition of PT-DNA, we determined the crystal structure of the Streptomyces coelicolor FL ScoMcrA to 3.15 Å resolution (PDB accession number 5ZMM) using the single-wavelength anomalous dispersion (SAD) method with a selenomethionine (SeMet) derivative (Fig. 1b–d, Supplementary Fig. 1, Supplementary Table 1)
Our study reveals that in addition to an irregular loop that binds in the major groove of GPSGCC, a highly conserved cavity is employed in the sulfur-binding domain (SBD) domain to recognize the phosphorothioate in PT-DNA, with both hydrophobic and electrostatic interactions
Summary
There have been very few reports on protein domains that recognize sulfur. We present the crystal structure of the sulfur-binding domain (SBD) from the DNA phosphorothioation (PT)-dependent restriction endonuclease ScoMcrA. We show that three of these homologs bind PT-DNA in vitro and restrict PT-DNA gene transfer in vivo. These results show that SBD-like PT-DNA readers exist widely in prokaryotes. The reported biological functions of DNA PT modification include conferring resistance to oxidation to the host bacteria[9,10,11,12,13], restricting gene transfer among different bacteria[14,15], and influencing the global transcriptional response[16,17], among others. As either modification is sufficient to elicit cleavage, it has been postulated that more than one recognition domain of ScoMcrA could be involved in the discrimination of modified PTDNA from normal DNA30
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