Hammerhead Ribozyme Structure
David Marcey an Cora Crenwelge
© David Marcey 2021

I. Introduction
II. Structure of the RNA-DNA Hammerhead Ribozyme
III. Structure of an All-RNA Hammerhead Ribozyme

IV. References

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I. Introduction

Hammerhead ribozymes are small, catalytic RNAs that undergo self-cleavage of their own backbone to produce two RNA products. All hammerhead ribozymes contain three base-paired stems and a highly conserved core of residues required for cleavage. The cleavage reaction proceeds by an attack of a 2' hydroxyl oxygen of a catalytic site cytosine on the phosphorus atom attached to the 3' carbon of the same residue. This breaks the sugar phosphate backbone and produces a 2', 3' cyclic phosphate. As for protein ribonucleases, a metal ion bound in the active site (Mg++) stabilizes the ionized form of the 2' hydroxyl oxygen, promoting the catalytic attack.

The wishbone-shaped structure at left is is a hammerhead ribozyme in which a portion of the RNA has been substituted with a stretch of DNA that acts as an inhibitor of catalysis, due to the absence of 2' hydroxyl groups in DNA (Pley, et al., 1994). The structure was the first described for a hammerhead ribozyme.

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II. Structure of the RNA-DNA Hammerhead Ribozyme

Stem I is formed by base pairing of complementary nucleotides through Watson-Crick hydrogen bonding. The 5' phosphate and 3' oxygen atoms mark the beginning and end of the ribozyme and are juxtaposed at the top of Stem I.

Stem II lies opposite Stem I in the other branch of the "wishbone." It also is formed by standard Watson-Crick base pairing and is capped by a four base loop at the top of the branch.

Stem III is formed by Watson-Crick base pairing and forms the base of the "wishbone." In a standard ribozyme, formed by a continuous RNA molecule instead of an RNA-DNA, two-chain hybrid, the nucleotides at the base of Stem III are connected by a short loop (see below).

The base sequences of the three stems can vary, depending on the hammerhead ribozyme under consideration. The only sequence restriction is that Watson-Crick base complementarity is maintained so that stem hydrogen bonding can form. The central core sequence, however, is highly conserved between ribozymes and is essential for ribozyme catalytic activity. The core contains two domains.

Domain 1 of the core comprises a CUGA sequence, the uridine turn, that follows Stem I.

Domain 2 is formed by non-Watson Crick base pairing, and connects Stem II and Stem III.

C17 is the conserved residue at which strand cleavage takes place (see Introduction).


 

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III. Structure of an All-RNA Hammerhead Ribozyme

Scott, et al. (1995) succeeded in determining the structure of a hammerhead ribozyme that comprises an RNA enzyme strand complexed with an RNA substrate strand, shown at left.

The substrate is not cleaved because the cleavage site cytosine bears a 2' methoxyl group instead of the reactive 2' hydoxyl (see Introduction).

Despite different crystallization conditions, loop position, and phosphate backbone connectivity, the structure of the all-RNA ribozyme is remarkably similar to the RNA-DNA hybrid discussed above, with the same stems (I, II, III), conserved core domains (CUGA and Domain 2), and catalytic C17. This suggests that these conformations likely represent the genuine structure of hammerhead ribozymes in solution.


 

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IV. References

Pley, H. W., Flaherty, K. M., McKay, D. B.: Three-dimensional structure of a hammerhead ribozyme. Nature 372: 68-74 (1994).

Scott, W.G., Finch, J.T., Klug, A.: The Crystal Structure of an All-RNA Hammerhaead Ribozyme: A Proposed Mechanism for RNA Catalytic Cleavage. Cell 81: 991-1002 (1995).


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