Biochemistry, Vol. 2 - A. Lehninger 1985

Bioenergetics and Metabolism
Metabolism: An Overview
Isotope labeling is a highly effective method for studying metabolism.

Another powerful method that makes it possible to trace the general course of a given metabolic pathway involves The Use of specific isotopes introduced as labels into a particular metabolite (Table 13-1). For example, the radioactive carbon isotope ¹⁴С (with an average atomic mass of 12.01) is frequently introduced into organic molecules as a label. Chemically, a labeled molecule is indistinguishable from a normal, unlabeled molecule, but its radioactivity allows it to be easily detected and its fate followed. For this purpose, one can, for instance, synthesize acetic acid in which the carbon of the carboxyl group is enriched with the radioactive isotope ¹⁴С. Under normal conditions, this isotope is present in the carbon compounds of the biosphere and geosphere in extremely small and constant concentrations. By feeding an animal ¹⁴С-acetate, one can trace the metabolic fate of this compound. We will then find, for example, that the exhaled CO₂ of the animal contains ¹⁴С, which demonstrates that a certain portion of the acetate undergoes metabolic transformations whereby the carbon of its carboxyl group is incorporated into CO₂. If palmitic acid is subsequently isolated from the animal's Liver Lipids, it is also found to contain ¹⁴С; consequently, the carboxyl carbon of acetate serves as a biosynthetic precursor of palmitic acid. Experiments involving the chemical Cleavage of such palmitic acid have further revealed that excess ¹⁴С is not characteristic of all carbon positions in its molecule, but only of positions at alternating carbon atoms starting from the carboxyl group (Fig. 13-21). If, however, an animal is fed acetate labeled with ¹⁴С exclusively in the methyl group, the labeled positions in the palmitic acid molecule will again be alternating carbon atoms, but this time counted from the α-carbon, or C-2. These observations led to the Conclusion that all carbon atoms of palmitic acid originate from acetate molecules and that during the synthesis of palmitic acid, the carbon skeletons of the acetate molecules are joined in a "HEAD-to-tail" fashion.

Class="center">Table 13-1. Some isotopes used as tracers

Fig. 13-21. Application of a radioactive carbon isotope to trace the metabolic fate of the carbon atom in the carboxyl group of acetate. A significant portion of the radioactive carbon from the labeled acetate is found in the exhaled CO₂, but a fairly large amount of it also ends up in the palmitic acid of liver lipids. In the palmitic acid molecule, only the odd-numbered carbon atoms (counted from the carboxyl group; shown in red) are labeled, indicating that palmitic acid is formed by the joining of eight acetate molecules in a "head-to-tail" manner.

The isotope tracer method is also used to determine the rates of turnover processes in the intact Organism. One of the most important findings obtained using this highly powerful method is the discovery that macromolecular components of Cells and Tissues undergo continuous metabolic turnover; in other words, the intracellular concentration of these components at any given moment is dynamic, representing the net result of continuous and balanced rates of Biosynthesis AND DEGRADATION. For example, isotope labeling studies established that the half-life of rat liver Proteins is 5–6 days (Table 13-2). At the same time, it has been shown that the turnover of Skeletal Muscle or Brain proteins is much slower.

We are indebted precisely to the isotope tracer method for A number of critical observations regarding METABOLISM.

Table 13-2. Metabolic turnover of some rat tissue components (based on an early study using radioactive carbon)