| Autor: |
Walker, John M., Turner, Philip C., De Young, Mary Beth, Siwkowski, Andrew, Hampel, Arnold |
| Zdroj: |
Ribozyme Protocols; 1997, p209-220, 12p |
| Abstrakt: |
Ribozymes have been successfully designed to downregulate gene expression in vivo. To enhance the probability of success in vivo, the investigator should have available the ribozyme with the highest catalytic activity feasible. This can only be determined by in vitro assays for catalytic efficiency and engineering the ribozyme appropriately to optimize this catalytic efficiency. Catalytic efficiency is kcat/km, where kcat is the turnover number of the reaction and km the true Michaelis constant. The kcat and Km values are best obtained individually by measuring catalytic activity in reactions where ribozyme is limiting and a range of excess substrate concentrations are used such that the ribozyme turns over (1). Functionally, km is the substrate concentration required to achieve half-maximum reaction velocity (Vmax), and the turnover number kcat is obtained by dividing the Vmax by the ribozyme concentration (Vmax/[Rz]) since Vmax = kcat[Rz]. The km is a combination of the individual rate constants comprising the overall reaction and, as such, does not reflect the rate of any given reaction step. [ABSTRACT FROM AUTHOR] |
| Databáze: |
Supplemental Index |
| Externí odkaz: |
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