Hammerhead ribozyme catalysis

Case Study Role of Divalent Metal Ions in Hammerhead Ribozyme Catalysis... 

Bertrand, E.L. and Rossi, J.J. (1994) Facilitation of hammerhead ribozyme catalysis by the nucleocapsid protein of HIV-1 and the heterogeneous nuclear ribonucleoprotein Al. EMBO J., 13,2904-2912. 

Herschlag, D., Khosla, M., Tsuchihashi, Z. and Karpel, R.L. (1994) An RNA chaperone activity of non-specific RNA binding proteins in hammerhead ribozyme catalysis. EMBOJ., 13, 2913-2924. 

Lee, T.-S., Silva-Lopez, C., Martick, M., Scott, W.G., York, D.M. Insight into the role of Mg2+ in hammerhead ribozyme catalysis from x-ray crystallography and molecular dynamics simulation. J. Chem. Theory Comput. 2007, 3, 325-7. 

Many researchers refer to stems 1, 2, and 3 using their Roman numeral equivalents—that is, stems I, II, and III. These motifs are also denoted as helices I, II, and III. It should be noted at the beginning of this hammerhead ribozyme discussion that structure-function relationships, the role of various nucleobases, metal ion participation in catalysis, and other features of the system have not been completely delineated and in some cases remain controversial. Globally, the hammerhead fold appears to be similar in both solution and solid-state studies. In solution, however, the central core of the hammerhead construct appears to be highly dynamic. This may account for different experimental results among the analytical techniques used in solution and certainly explains some distinct differences seen between solution and solid-state (X-ray crystallographic) structures. 

Additional interactions and rearrangements in the transition state with other rescuing bases may take place because it is known from crystal structures that substrate atoms are not in line for nucleophilic attack in the hammerhead ribozyme (at least not in the published crystal structures). Also, a metal ion located -20 A away from the catalytic site was shown to be crucial for catalysis. This same metal ion appeared likely to take on an additional ligand in the transition state, suggesting that conformational changes had to take place before catalysis. ... 

Fig. 1A-F The two-dimensional structures of various ribozymes. The ribozyme or intron portion is printed in black. The substrate or exon portion is printed in gray. Arrows indicate sites of cleavage by ribozymes A (left) the two-dimensional structure of a hammerhead ribozyme and its substrate. Outlined letters are conserved bases that are involved in catalysis right) The y-shaped structure of the hammerhead ribozyme-sub-strate complex B-F the two-dimensional structures of a hairpin ribozyme, the genomic HDV ribozyme, a group I ribozyme from Tetrahymena, a group II ribozyme from S. cer-evisiae (aiy5), and the ribozyme of RNase P from E. coli...

Specificity of conventional protein enzymes is provided by precise molecular fit. The mutual recognition of an enzyme and is substrate is the result of various intermolecular forces which are almost always strongly dominated by hydrophobic interaction. In contrast, specificity of catalytic RNAs is provided by base pairing (see for example the hammerhead ribozyme in Figure 1) and to a lesser extent by tertiary interactions. Both are the results of hydrogen bond specificity. Metal ions too, in particular Mg2+, are often involved in RNA structure formation and catalysis. Catalytic action of RNA on RNA is exercised in the cofolded complexes of ribozyme and substrate. Since the formation of a ribozyme s catalytic center which operates on another RNA molecule requires sequence complementarity in parts of the substrate, ribozyme specificity is thus predominantly reflected by the sequence and not by the three-dimensional structure of the isolated substrate. 

Figure 1. Catalysis and template action of RNA and proteins. Catalytic action of one RNA molecule on another one is shown in the simplest case, the "hammerhead ribozyme." The substrate is a tridecanucleotide forming two double-helical stacks together with the ribozyme (n = 34) in the confolded complex. Tertiary interactions determine the detailed structure of the hammerhead ribozyme complex and are important for the enzymatic reaction cleaving one of the two linkages between the two stacks. Substrate specificity of ribozyme catalysis is caused by secondary structure in the cofolded complex between substrate and catalyst. Autocatalytic replication of oligonucleotide and nucleic acid is based on G = C and A = U complementarity in the hydrogen bonded complexes of nucleotides forming a Watson-Crick type double helix. Gunter von Kiedrowski s experi-...

Interestingly, specific M +-nucleic acid interactions determined in solution for the hairpin ribozyme by spectroscopic and chemical interference methods were not later found in their X-ray crystal stmctures. Even more confusing were results of X-ray crystallography " and initial solution experiments, which both determined up to two, presumably functionally important Mg + ions in the catalytic center of the hammerhead ribozyme. However, as demonstrated recently in biochemical experiments in solution, neither of these ribozymes require either inner- or outer-sphere interactions with particular metal ions for catalysis. ... 

Keywords RNA catalysis molecular simulation hammerhead ribozyme LI ligase ribozyme Mg2 ions... 

Han, J., Burke, J.M. Model for general acid-base catalysis by the hammerhead ribozyme pH-activity relationships of G8 and G12 variants at the putative active site. Biochemistry 2005, 44, 7864-70. 

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