Single-hit kinetics

RNA interaction and therefore cannot be recommended for probing experiments (e.g. RNase SI and RNase U2 have a pH optimum of 4.5 which prevents or destabilises protein-RNA complex formation). RNase A has a high affinity for a pyrimidine-adenosine stretch (particularly UA), so it can therefore be difficult to obtain single-hit kinetics (except for the UA sequence). Furthermore, RNase A exhibits an endogenous helix unfolding property which makes structure assignment difficult. RNases such as RNase CL3 and a-sarcin are inhibited by Mg2+ which is required for the stability of many complexes and also for proper folding of RNA. 

Fig. 4.13. Flow chart of the protein footprinting technique. Purified recombinant protein, which has been radioactively labelled at the N- or C-terminal, is cleaved by proteinases or chemicals with single-hit kinetics. The substrate is kept at native conditions in uncomplexed and complexed form. The cleavage products are analysed in high resolution SDS gels.

It is not known whether one or both of the two types of AAV virions is individually capable of initiating a productive infection in the presence of adenovirus or whether both types of AAV particles are required. Titration studies of AAV infectivity in the presence of excess adenovirus have suggested single-hit kinetics for infection this would imply that it is not essential for both types of co-infect a cell (Blacklow et al., 1967). Attempts to productively infect cells with physically separated particles (Berns and Adler, 1972) have been made but the data have not been of sufficient precision to resolve this problem (Hoggan and Berns, unpublished data). Purified AAV DNA has been shown to be infectious in the presence of helper adenovirus (Hoggan et al., 1968). The infectivity of the DNA was increased by the addition of... 

To substantiate this concept, the present set of experiments should be complemented by analogous data from transfection experiments with phage DNAs showing single-hit kinetics due to transfection with monomeric molecules. The model would predict that in these cases transfection crosses would not yield results which are basically different from normal phage crosses. Unfortunately, 029 can not be used for this purpose. 029 DNA molecules show a tendency to aggregate (Hirokawa, unpublished). Studies of genetic recombination in the SPO 2 system are presently under way in our laboratory. 

Mention was made in Section XVIII-2E of programmed desorption this technique gives specific information about both the adsorption and the desorption of specific molecular states, at least when applied to single-crystal surfaces. The kinetic theory involved is essentially that used in Section XVI-3A. It will be recalled that the adsorption rate was there taken to be simply the rate at which molecules from the gas phase would strike a site area times the fraction of unoccupied sites. If the adsorption is activated, the fraction of molecules hitting and sticking that can proceed to a chemisorbed state is given by exp(-E /RT). The adsorption rate constant of Eq. XVII-13 becomes... 

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