Structural insight into the novel <i>Thermus thermophilus</i> SPOUT methyltransferase RlmR catalysing Um2552 formation in the 23S rRNA A-loop: a case of convergent evolution

The A-loop of the 23S ribosomal RNA is a critical region of the ribosome involved in stabilizing the CCA-end of A-site-bound transfer RNA. Within this loop, nucleotide U2552 is frequently 2'-O-methylated (Um2552) in various organisms belonging to the three domains of life. Until now, two enzymatic systems are known to modify this position, relying on either a Rossmann fold-like methyltransferase (RFM) or a small RNA-guided system. Here, we report the identification of a third system involved in Um2552 formation, consisting of a methyltransferase of the SPOUT (SpoU-TrmD) superfamily encoded by the ttc1712 open reading frame of Thermus thermophilus, herein renamed RlmR. In Escherichia coli and human mitochondria, the absence of the RFM enzyme responsible for Um2552 formation is known to cause severe defects in ribogenesis and ribosome function. In contrast, no comparable effect was observed upon ttc1712 gene invalidation in T. thermophilus. We also report the high-resolution crystal structure of RlmR in complex with a 59-mer substrate RNA. The structure highlights significant conformational rearrangements of the A-loop and provides a new insight into the catalytic mechanism, revealing structural features that may be generalized to other SpoU methyltransferases.