The Enzymology of Control by Feedback Inhibition

One way in which an organism controls its biosynthetic path- ways is by feedback inhibition. This control adjusts the rate of production of the end products of pathways, such as amino acids and nucleotides, to the rate of synthesis of macromolecules, such as proteins and nucleic acids. Control is imposed by the end product itself in a simple and rapid manner: the end product stops production of more of itself by inhibiting the activity of the first enzyme unique to its pathway. Consequently, if an end product is not continually removed into macromolecules, additional end product is not made. The organism benefits from feedback inhibition, because it avoids wasting resources of carbon, nitrogen, and energy on end products which are not used. Certainly it is a widespread and effective metabolic con- trol. Many biosynthetic pathways in bacteria have feedback inhibition, and a few instances are also known for higher or- ganisms (1). In this article, a feedback inhibition is investigated to learn more about its enzymological mechanism. Until now, feedback inhibition has been studied at a metabolic level where one us- ually asks which end product inhibits which enzyme. The question now asked is, how does the end product of a pathway inhibit the first enzyme? This question is justified because many known feedback inhibitions appear competitive, as if in- hibition results from a competition of the inhibitor and substrate for the same groups of the active site on the first enzyme. To the enzymologist, competition is wholly unexpected because the end product and substrate often differ greatly in size, shape, and charge, in contrast with classical competitors (2). How can there be competition when a site specific for the substrate seems unsuited for also binding the end product? The problem of the mechanism of inhibition was approached by using the first enzyme unique to pyrimidine biosynthesis in Escherichiu coli. This enzyme is aspartate transcarbamylase, which catalyzes the reaction shown in Fig. 1. It is known that in the bacterium the activity of aspartate transcarbamylase is closely controlled by an end product, probably a cytosine de- rivative, and that inhibition of this enzyme is important for controlling the whole pathway (3). Aspartate transcarbamylase is especially suited for enzymological study because it can be obtained easily in a highly purified state (4). It is inhibited by a nucleotide end product which competes with a structurally lunrelated substrate, an amino acid. J Experiments to be described have two purposes: first, to find * This work was aided bv support from United States Public