Follow-up experiments

To provide confirmation of the results of the experiment by follow-up experiments. 

Follow-up experiments with the similar Klebsiella pneumoniae enzyme (66, 67) also showed that the pH profiles for CatPx are quite different from those for classical catalases. The latter s catalatic activities are essentially pH-independent from pH 5 to 10 (6S) CatPx of Klebsiella showed a sharp pH optimum between pH 6 and 7 (66). A similar eukaryotic fungal CatPx (69) was also characterized by a sharp pH sensitivity and saturation kinetics K 3.4 mM) and, like the bac-... 

In follow-up experiments, Gobas et al. (1989) found that brominated biphenyls which have minimal internal cross sections somewhat greater than 0.95 nm were available for uptake in fish via the gills. In addition, Muir et al. (1986) and Muir and Yarechewski (1988) reported that octachlorodibenzo-p-dioxin, which has a minimal internal cross section greater than 0.95 nm, was present in internal tissues of trout exposed under field conditions. 

Initial experiments with DEBS 1-TE showed that only the (2S)-isomer of methylmalonyl-CoA is used to generate both stereocenters (d and l) in the trike-tide lactone (Fig. 13) [36]. In follow-up experiments, DEBS 1-TE was incubated in the presence of starter unit, NADPH, and deuterium-labeled (2S)-methylmalo-nyl-CoA [42], Use of the isotopically labeled substrate allowed discrimination between possible mechanisms by which the synthase could generate two methyl stereochemistries from a precursor with a single methyl configuration (Fig. 14), because the deuterium labeling pattern in the resulting lactone was characteristic... 

If necessary, subsequent experiments can then determine irreversibility and potency (e.g., Ki and jnact) f°r metabolism-dependent inhibitors, as well as the potential for covalent binding. This general experimental strategy, including follow-up experiments, is illustrated in Figure 5. The design of experiments to determine Kt and /clnact will be covered in more detail in section II.C.7.C. 

Figure 5 Flowchart for initial and possible follow-up CYP inhibition studies. The first box represents the IC50 experiment with and without a 30-minute preincubation with NADPH (the highest concentration of test article is also preincubated for 30 minutes without NADPH). Remaining boxes depict possible outcomes and follow-up experiments. Abbreviations CYP, cytochrome P450 IC50, concentration of inhibition causing 50% inhibition.

Identification of the endogenous substrate(s) of an enzyme has an immediate effect on the biochemical characterization of the protein, but it can also have a broader impact on downstream metabolic and signaling studies. For example, the discovery of NATs naturally implies the presence of a previously unappreciated biosynthetic pathway responsible for the production of these compounds. Indeed, follow-up experiments confirmed the existence of a membrane-bound enzyme activity in the liver and kidney capable of producing NATs from acyl-CoAs and taurines [29] (Fig. 7a). 

Circumstantial evidence was observed in the post-bombardment surface morphology of gallium samples that were exposed to deuterium plasma and then rapidly cooled [20]. Small voids were observed in the sample surface that may have resulted from the coalescence of bubbles during the sample cool down period. Consistent with this view was the fact that the amount of deuterium retained in the samples was independent of the sample exposure conditions. Follow-up experiments are being performed to determine whether gallium samples exposed to deuterium plasma at elevated temperature (around... 

It should be noted that these measurements report the relative abrmdance of each protein in the ceU extracts. Isotope ratios and comparative densitometry do not distinguish changes in expression, regulation, or degradation. Follow-up experiments are required to make these distinctions. 

If a structure has passed all the requirements thus far, one needs to determine if it can be derived from a plausible synthetic route. In other words, is it consistent with the reaction pathway and chemical entities present in the reaction vessel If additional structural information is required, it is necessary at this point to identify appropriate follow-up experiments. For example, if the stereochemistry at a chiral center is critical to the understanding of the system, nOe experiments may be useful to investigate spatial relationships. To improve the confidence level in a proposed structure, it may be necessary to synthesize the proposed structure using a well-established synthetic route and then repeat the NMR characterization on the synthesized compound. Comparing the proton and carbon spectra of the unknown with the known compound should confirm consistency of structure. Finally, the team should draw conclusions based on the identified structure, plausible synthetic routes, and ways to prevent its formation. 

In order to apply a statistical interpretation, we need replication of the identified types of land. Replicate areas do not have to be contiguous they could be in separate fields but at a similar level, for example. The null hypothesis is that the soils from the different types of land are not different in available P content. A follow-up experiment could determine the P contents of vegetation associated with the soil sites. 

This is the same task presented to the four groups of individuals as part of the validation of the situations, as described in chapter 3, and is also the same task used in the follow-up experiment described earlier in this chapter. 

In a follow-up experiment three volunteers were placed on this same very-low-folate diet and supplemented with 25, 50, and 100 p.g of folate daily (H17). After 1 month there was a fall in the serum folate level from 10.3 to... 

---