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Publication Abstract from PubMed

RtcB is an atypical RNA ligase that joins either 2',3'-cyclic phosphate or 3'-phosphate termini to 5'-hydroxyl termini. In contrast to typical RNA ligases, which rely on ATP and Mg(II), catalysis by RtcB is dependent on GTP and Mn(II) with ligation proceeding through a covalent RtcB-histidine-GMP intermediate. Here, we present three structures of Pyrococcus horikoshii RtcB complexes that capture snapshots along the entire guanylylation pathway. These structures show that prior to binding GTP, a single manganese ion (Mn1) is bound to RtcB. To capture the step immediately preceding RtcB guanylylation, we determined a structure of RtcB in complex with Mn(II) and the unreactive GTP analogue guanosine 5'-(alpha-thio)triphosphate (GTPalphaS). This structure shows that Mn1 is poised to stabilize the pentavalent transition state of guanylylation while a second manganese ion (Mn2) is coordinated to a nonbridging oxygen of the gamma-phosphoryl group. The pyrophosphate leaving group of GTPalphaS is oriented apically to His404 with the epsilon-nitrogen poised for in-line attack on the alpha-phosphorus atom. The structure of RtcB in complex with GTPalphaS also reveals the network of hydrogen bonds that recognize GTP and illuminates the significant conformational changes that accompany the binding of this cofactor. Finally, a structure of the enzymic histidine-GMP intermediate depicts the end of the guanylylation pathway. The ensuing molecular description of the RtcB guanylylation pathway shows that RtcB and classical ATP- and Mg(II)-dependent nucleic acid ligases have converged upon a similar two-metal mechanism for formation of the nucleotidylated enzyme intermediate.

Structures of the Noncanonical RNA Ligase RtcB Reveal the Mechanism of Histidine Guanylylation.,Desai KK, Bingman CA, Phillips GN Jr, Raines RT Biochemistry. 2013 Apr 16;52(15):2518-25. doi: 10.1021/bi4002375. Epub 2013 Apr, 5. PMID:23560983^[1]

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.