Crystal structure of an ASCH protein from Zymomonas mobilis and its ribonuclease activity specific for single-stranded RNA.

Activating signal cointegrator-1 homology (ASCH) domains were initially reported in human as a part of the ASC-1 transcriptional regulator, a component of a putative RNA-interacting protein complex; their presence has now been confirmed in a wide range of organisms. Here, we have determined the trigonal and monoclinic crystal structures of an ASCH domain-containing protein from Zymomonas mobilis (ZmASCH), and analyzed the structural determinants of its nucleic acid processing activity. The protein has a central β-barrel structure with several nearby α-helices. Positively charged surface patches form a cleft that runs through the pocket formed between the β-barrel and the surrounding α-helices. We further demonstrate by means of in vitro assays that ZmASCH binds nucleic acids, and degrades single-stranded RNAs in a magnesium ion-dependent manner with a cleavage preference for the phosphodiester bond between the pyrimidine and adenine nucleotides. ZmASCH also removes a nucleotide at the 5'-end. Mutagenesis studies, guided by molecular dynamics simulations, confirmed that three residues (Tyr47, Lys53, and Ser128) situated in the cleft contribute to nucleic acid-binding and RNA cleavage activities. These structural and biochemical studies imply that prokaryotic ASCH may function to control the cellular RNA amount.