Hammerhead Ribozymes

Abstract

The hammerhead ribozyme is an RNA‐cleaving, RNA enzyme, derived from single‐stranded RNA viroids, and virusoids, which infect flowering plants. Engineering of hammerhead ribozymes allows these RNA catalysts to be utilized for cleaving virtually any RNA target of choice, thus making them useful tools for both genetic and therapeutic applications.

Keywords: ribonucleic acid; RNA; gene therapy; catalysis; site‐specific cleavage

Figure 1.

The hammerhead ribozyme involved in base pairing with a target RNA. N represents any of the four nucleotides, A (adenosine), C (cytosine), G (guanosine) and U (uridine). The H represents A, C or U; Y = C or U, and P = A or G. The numbers define key bases in the catalytic core of the ribozyme. The G5 is cited as an example of a key position in the catalytic core that cannot be altered without crippling ribozyme catalytic activity. The U4 nucleotide is the U of the U turn.

Figure 2.

(a) The hammerhead consensus sequence in the context of the three‐dimensional structure. The conventions used are the same as those of Figure . In this representation, the ribozyme is shown in its cis configuration, as it might appear in the native substrate of a viroid or virusoid single‐stranded RNA. The U turn is indicated. Noncannonical (non‐Watson–Crick) base interactions are indicated by heavy bars between the bases, whereas Watson–Crick‐type base pairings are indicated by the double dashes between bases. (b) The hammerhead ribozyme mechanism. The ribozyme coordinates Mg2+ in the proper geometry to stabilize the trigonal bipyramid intermediate formed by attack of the 2′‐OH on the phosphate. The end products are a 2′,3′ cyclic phosphate and a 5′‐OH. For 3D structures, visit the following website [http://www.tulane.edu/∼biochem/nolan/lectures/rna/frames/hambtx.htm].

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References

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Further Reading

Kearney P, Wright LA, Milliken S and Biggs JC (1995) Improved specificity of ribozyme‐mediated cleavage of bcr‐abl mRNA. Experimental Hematology 23: 986–989.

Koizumi M, Kamiya H and Ohtsuka E (1992) Ribozymes designed to inhibit transformation of NIH3T3 cells by the activated c‐Ha‐ras gene. Gene 117: 179–184.

Morgan RA and Anderson WF (1993) Human gene therapy. Annual Review of Biochemistry 62: 191–217.

Rossi JJ (1994) Practical ribozymes. Making ribozymes work in cells. Current Biology 4: 469–471.

Rossi JJ (1995) Controlled, targeted, intracellular expression of ribozymes: progress and problems. Trends in Biotechnology 13: 301–306.

Scott WG, Murray JB, Arnold JRP, Stoddard BL and Klug A (1996) Capturing the structure of a catalytic RNA intermediate: the hammerhead ribozyme. Science 274: 2065–2069.

Shimayama T, Nishikawa S and Taira K (1995) Generality of the NUX rule: kinetic analysis of the results of systematic mutations in the trinucleotide at the cleavage site of hammerhead ribozymes. Biochemistry 34: 3649–3654.

Sullenger BA and Cech TR (1993) Tethering ribozymes to a retroviral packaging signal for destruction of viral RNA. Science 262: 1566–1569.

Vaish NK, Kore AR and Eckstein F (1998) Recent developments in the hammerhead ribozyme field. Nucleic Acids Research 26: 5237– 5242.

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Rossi, John R(Apr 2001) Hammerhead Ribozymes. In: eLS. John Wiley & Sons Ltd, Chichester. http://www.els.net [doi: 10.1038/npg.els.0003133]