Ribozyme mediated

As the concentration of NaCl is increased after ribozyme activity has reached a minimum, the activity of the ribozyme is restored (Fig. 8). This rescue cannot be explained in terms of the number of bound Mn ions since the number does not increase at higher concentrations of NaCl, according to Horton et al., as described above. It is likely that, at higher concentrations, Na ions can work to fold the complex into the active structure and, as a result, more efficient Mn ions, even at limited concentrations, can work as a catalyst (see below for details). More efficient Mn ions can work as a catalyst even at a limited concentrations. As we would anticipate from the structural role and inefficient catalytic activity of Na ions, several groups have reported that ribozyme-mediated cleavage reactions can occur even in the absence of divalent metal ions provided that the concentration of monovalent ions, such as Na" ions, is extremely high [64-66]. 

Figure 4.3 Chimeric hammerhead ribozyme structure. The ribozyme structure was computer modeled after the work of Pley et al. (1994). The yellow ribbon (backbone) and bases depict the catalytic core of the ribozyme, while the green ribbon (backbone) and bases depict the binding arms, which in this case are DNA from the work of Taylor etal. (1992). The magenta bases and ribbon (backbone) are the target RNA. The orange ribbon depicts the backbone of stem II, which can be minimally reduced to four propane diols and one G C base pair (J.Rossi and P.Swiderski, unpublished data). The blue ribbon represents the target backbone at the site of ribozyme-mediated cleavage, (see Color Plate 2)...

Ribozyme-mediated target validation can also be used to identify specific member(s) of a protein family involved in a specific phenotype. Carefully designed ribozymes can selectively knock down expression of each protein in a gene family. 

Table 4.1 Partial list of successful ribozyme mediated target mRNA down-regulation Target Effect Reference...

Efrat, S., Leiser, M., Wu, Y.J., Fusco-DeMane, D., Emran, O.A., Surana, M. et al. (1994) Ribozyme-mediated attenuation of pancreatic beta-cell glucokinase expression in transgenic mice results in impaired glucose-induced insulin secretion. Proc. Natl. Acad. Sci. USA, 91, 2051-2055. 

Gu, J.L., Pei, H., Thomas, L., Nadler, J.L., Rossi, J.J., Lanting, L. and Natarajan, R. (2001) Ribozyme-mediated inhibition of rat leukocyte-type 12-lipoxygenase prevents intimal hyperplasia in balloon-injured rat carotid arteries. Circulation, 103, 1446-1452. 

Michienzi, A., Cagnon, L., Bahner, I. and Rossi, J.J. (2000) Ribozyme-mediated inhibition of HIV 1 suggests nucleolar trafficking of HIV-1 RNA. Proc. Natl. Acad. Sci. USA, 97, 8955-8960. 

Scherr, M., Reed, M., Huang, C.F., Riggs, A.D. and Rossi, J.J. (2000) Oligonucleotide scanning of native mRNAs in extracts predicts intracellular ribozyme efficiency ribozyme-mediated reduction of the murine DNA... 

Snyder, D.S., Wu, Y., Wang, J.L., Rossi, J.J., Swiderski, P., Kaplan, B.E. and Forman, S.J. (1993) Ribozyme-mediated inhibition of bcr-abl gene expression in a Philadelphia chromosome-positive cell line. Blood, 82,600-605. 

Sullenger, B.A. and Cech, T.R. (1994) Ribozyme-mediated repair of defective mRNA by targeted, trans-splicing. Nature, 371, 619-622. 

D.-M. Zhou, K. Taira, The Hydrolysis of RNA From Theoretical Calculations to the Hammerhead Ribozyme-Mediated Cleavage of RNA , Chem. Rev., 98,991 (1998)... 

Kanazawa Y, Ohkawa K, Ueda K, Mita E, Takehara T, et al. 1996. Hammerhead ribozyme-mediated inhibition of telomerase activity in extracts of human hepatocellular carcinoma cells. Biochem. Bio-phys. Res. Commun. 225 570-76... 

Weinberg, M., Passman, M., Kew, M., Arbuthnot, P. Hammerhead ribozyme-mediated inhibition of hepatitis b virus x gene expression in cultured cells. J. Hepatol. 2000, 33(1), 142-51. 

Figure 3 (See color insert) Scheme of ribozyme-mediated cleavage of target mRNA molecules. The ribozyme (in red) is designed to target mRNA sequence centered on the desired cleavage site (asterisk). It cleaves this mRNA sequence specifically.

Figure 11.3 Scheme of ribozyme-mediated cleavage of target mRNA molecules. (See p. 165)... 

Sun L.-Q., Warrilow D., Wang L., Witherington C., Macpherson J., Symonds G. (1994) Ribozyme-mediated suppression of Moloney murine leukemia virus and human immunodeficiency virus type I replication in permissive cell lines. Proc Natl. Acad. Sci. USA 91 9715. 

Heidemeich, O., Eckstein, F. (1992). Hammerhead ribozyme-mediated cleavage of the long terminal repeat RNA of human immunodeficiency virus type 1. J Biol Chem 267, 1904-1909. 

Clark, D. R., Zacharias, W., Panaitescu, L., and McGregor, W. G. (2003). Ribozyme-mediated REVl inhibition reduces the frequency of UV-induced mutations in the human HPRT gene. Nucleic Adds Res. 31, 4981-4988. 

An artificial ribozyme mimics this translation step of the ribosome. The spedlity of this selected ribozyme is based on the recognition of an adenosine moiety of the amino acid ester and allows the utilization of leucine- and phenylalanine- as weU as methionine-derivatized substrates. This tolerance for various amino acids indicates the possibility of selecting more general ribozymes for protein synthesis. Furthermore, a related ribozyme efficiently catalyses the synthesis of -30 different dipeptides from an aminoacyl-adenylate substrate. Ribozyme-mediated synthesis of uncoded peptides might have been an important step in the transition from a RNA to a peptide world before the anergence of the ribosome. ... 

In the Famulok laboratory, a ribozyme was isolated which promotes a reaction corresponding to the first step of the formation of dTMP from dUMP in proteic thymidylate synthases. This ribozyme mediates Michael-adduct formation at a Michael-acceptor substrate. The reaction is accelerated by a factor of nearly 10. The selected ribozyme could be engineered to act in an intermolecular reaction on a substrate tethered to an RNA oligomer. The danonstration of RNA catalysis of this reaction has bearings on the RNA world hypothesis, as well as implications for possible synthetic applications. ... 

Zhou D-M, Taira K (1998) The hydrolysis of RNA from theoretical calculations to the hammerhead ribozyme-mediated cleavage of RNA. Chem Rev 98 991-1026... 

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