The membrane-targeting-sequence motif is required for exhibition of recessive resurrection inEscherichia coliRNase E

Abstract

AbstractThe essential homo-tetrameric endoribonuclease RNase E ofEscherichia coliparticipates in global RNA turnover as well as stable RNA maturation. The protomer’s N-terminal half (residues 1-529) bears the catalytic, allosteric and tetramerization domains, including the critical active site residues D303 and D346. The C-terminal half (CTH, residues 530-1061) is dispensable for viability. We have previously described a phenomenon of recessive resurrection in RNase E that requires the CTH, wherein the wild-type homo-tetramer displays identical activity in vivo as a hetero-tetramer comprised of three catalytically dead subunits (with D303A/D346A substitutions) and one wild-type subunit. Here we show that recessive resurrection is exhibited even in dimeric RNase E with the CTH, and that it is dependent on presence of the membrane-targeting-sequence motif (residues 565-582). A single F575E substitution also abolished recessive resurrection, whereas other CTH motifs (such as those for binding ofRNAor of partner proteins) were dispensable. The phenomenon was independent of RNA 5’-monophosphate sensing by the enzyme. We propose that membrane-anchoring of RNase E renders it uniquely processive for endoribonucleolytic action, and that recessive resurrection and dominant negativity are alternative and mutually exclusive manifestations of, respectively, processive and distributive catalytic mechanisms in a homo-oligomeric enzyme.