Results and Discussion Structure Determination. The Pistol RNA construct used fo

Question

Results and Discussion Structure Determination. The Pistol RNA construct used for crys- tallization was derived from an extensive comparative genomic analysis of the previously identified environmental sample 27, ikely of bacterial construct contains two RNA strands annealed together: one being the enzyme strand and the other being a substrate strand (Fig, 1) (3) The substrate strand contains the Pistol self-deavage site, which is positioned between guanosine 10 (G10) and uridine 11 (U11) (Fig. IB). To trap the Pistol ribozyme in its precatalytic state for crystallization, we generated a Pistol RNA substrate Significance Based on the "RNA world' theory, ribozymes likely carried out biochemical reactions long before organisms evolved to use protein enzymes as biocatalysts. The continued discovery of new structures for small self-deaving ribozymes has shed light on conserved mechanisms in evolution, such as acid-base catalysis for self-deavage reaction. Here, we present the aystal structure of a newly discovered dass of self-deaving ribozymes called Pistol and how it likely uses the phosphoester transfer mecha- nism for self-cleavage. The results presented here suggest that Pistol uses an evolutionarily conserved cleavage mechanism that is like other self-cleaving ribozymes, sudh as Twister, Hammer head, Hairpin, and Hepatitis Delta Virus ribozymes. Author contributions: LAN- designed rescardu LAN. Performed research: TAS contrib- uted new reagentsanalyti took: LAN, uw. and T-A-s. analyzed data and LAN. and JAW. wrote the paper. The authors declare no conflict of interest This article is a PNAS Direct Submission. Data deposition The Drymtallography, atomic coordinates and structure factors have been sited in the Protein Data Bank, wwww. 'To whom correspondence may be addressed Emait jimin wangeyale edu or thomas. This artide contains upporting information online at wwww.pnas.orgiackupituppldok 107apnas 1611151 114DCsupplemental. PNAS January 31, 2017 vol. 114 no. 5

Explanation / Answer

The pistol RNA contains two RNA strands, enzymes and substrate strand, which is annealed together. The pistol cleavage site, present in substrate strand between G10 and U11. The data depicted in the experiments showed presence of Co-hexamine ion at the active portion. The structure is consisting of three helical stems, having Watson-Crick pairing (connected three loops), and one pseudoknot. It is reported that the 11bp constitute the coaxially stacked stem P1. there are 6 Watson-Crick pairs have observed in pseudoknot P1 while 4bp in P1 & P2 stems.

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