Overview of Nitrogen Metabolism:- Glutamine Synthetase Is a Primary Regulatory Point in Nitrogen Metabolism
The activity of glutamine synthetase is regulated in vir tually all organisms—not surprising, given its central metabolic role as an entry point for reduced nitrogen. In enteric bacteria such as E. coli, the regulation is un usually complex. The enzyme has 12 identical subunits of Mr 50,000 (Fig. 22–5) and is regulated both allosterically and by covalent modification. Alanine, glycine, and at least six end products of glutamine metabolism are allosteric inhibitors of the enzyme (Fig. 22–6). Each in hibitory alone produces only partial inhibition, but the effects of multiple inhibitors are more than additive, and all eight together virtually shut down the enzyme. This control mechanism provides a constant adjustment of glutamine levels to match immediate metabolic requirements.
FIGURE 22–5 Subunit structure of glutamine synthetase as determined by x-ray diffraction. (PDB ID 2GLS) (a) Side view. The 12 sub units are identical; they are differently colored to illustrate packing and placement. (b) Top view, showing active sites (green).
Superimposed on the allosteric regulation is inhibition by adenylylation of (addition of AMP to) Tyr397, located near the enzyme’s active site (Fig. 22–7). This co valent modification increases sensitivity to the allosteric inhibitors, and activity decreases as more subunits are adenylylated. Both adenylylation and deadenylylation are promoted by adenylyl transferase (AT in Fig. 22–7), part of a complex enzymatic cascade that responds to levels of glutamine, -ketoglutarate, ATP, and Pi. The activity of adenylyltransferase is modulated by binding to a regulatory protein called PII, and the activity of PII, in turn, is regulated by covalent modification (uridylylation), again at a Tyr residue. The adenylyl transferase complex with uridylylated PII (PII-UMP) stimulates deadenylylation, whereas the same complex
FIGURE 22–6 Allosteric regulation of glutamine synthetase. The en zyme undergoes cumulative regulation by six end products of glutamine metabolism. Alanine and glycine probably serve as indicators of the general status of amino acid metabolism in the cell.
FIGURE 22–7 Second level of regulation of glutamine synthetase: covalent modifications. (a) An adenylylated Tyr residue. (b) Cascade leading to adenylylation (inactivation) of glutamine synthetase. AT rep resents adenylyltransferase; UT, uridylyl transferase. Details of this cascade are discussed in the text.
with deuridylylated PII stimulates adenylylation of glut amine synthetase. Both uridylylation and deuridylylation of PII are brought about by a single enzyme, uridylyl transferase. Uridylylation is inhibited by binding of glutamine and Pi to uridylyltransferase and is stimulated by binding of α-ketoglutarate and ATP to PII. The regulation does not stop there. The uridylylated PII also mediates the activation of transcription of the gene encoding glutamine synthetase, thus increasing the cellular concentration of the enzyme; the deuridylylated PII brings about a decrease in transcription of the same gene. This mechanism involves an interaction of PII with additional proteins involved in gene regulation, of a type described in Chapter 28. The net result of this elaborate system of controls is a decrease in glutamine synthetase activity when glutamine levels are high, and an increase in activity when glutamine levels are low and α-ketoglutarate and ATP (substrates for the synthetase reaction) are available. The multiple layers of regulation permit a sensitive response in which glut amine synthesis is tailored to cellular needs.
