Protein turnover

Most proteins in the body are constantly being synthesized and then degraded (turned over), permitting the removal of abnormal or unneeded proteins. For many proteins, regulation of synthesis determines the concentration of protein in the cell, with protein degradation assuming a minor role. For other proteins, the rate of synthesis is constitutive (that is, essentially constant), and cellular levels of the protein are controlled by selective degradation.

1. Rate: In healthy adults, the total amount of protein in the body remains constant because the rate of protein synthesis is just sufficient to replace the protein that is degraded. This process, called protein turnover, leads to the hydrolysis and re-synthesis of 300–400 g of body protein each day.

The rate of protein turnover varies widely for individual proteins. Short lived proteins (for example, many regulatory proteins and mis-folded proteins) are rapidly degraded, having half-lives measured in minutes or hours. Long-lived proteins, with half-lives of days to weeks, constitute the majority of proteins in the cell. Structural proteins, such as collagen,  are metabolically stable and have half-lives measured in months or years.

2. Protein degradation: There are two major enzyme systems responsible for degrading proteins: the ATP-dependent ubiquitin (Ub)–proteasome system of the cytosol and the ATP-independent degradative enzyme system of the lysosomes. Proteasomes selectively degrade damaged or short-lived proteins. Lysosomes use acid hydrolases  to nonselectively degrade intracellular proteins (autophagy) and extracellular proteins (heterophagy), such as plasma proteins, that are taken into the cell by endocytosis.

a. Ubiquitin–proteasome system: Proteins selected for degradation by the cytosolic ubiquitin–proteasome system are first modified by the covalent attachment of Ub, a small, globular, nonenzymic protein that is highly conserved across eukaryotic species. Ubiquitination of the target substrate occurs through isopeptide linkage of the α-carboxyl group of the C-terminal glycine of Ub to the ε-amino group of a lysine in the protein substrate by a three-step, enzyme-catalyzed, ATP dependent process. [Note: Enzyme 1 (E1, an activating enzyme)  activates Ub, which is then transferred to E2 (a conjugating enzyme).

E3 (a ligase) identifies the protein to be degraded and interacts with E2-Ub. There are many more E3 proteins than there are E1 or E2.]  The consecutive addition of four or more Ub molecules to the target protein generates a polyubiquitin chain. Proteins tagged with Ub chains are recognized by a large, barrel-shaped, macromolecular,  proteolytic complex called a proteasome (Fig. 1). The proteasome unfolds, de-ubiquitinates, and cuts the target protein into fragments that are then further degraded by cytosolic proteases to amino acids, which enter the amino acid pool. The Ub is recycled. It is noteworthy that the selective degradation of proteins by the ubiquitin–proteosome complex  (unlike simple hydrolysis by proteolytic enzymes) requires ATP hydrolysis.

Figure 1:  The ubiquitin–proteasome degradation pathway of proteins. AMP = adenosine monophosphate; PPi = pyrophosphate.

b. Degradation signals: Because proteins have different half-lives, it is clear that protein degradation cannot be random but, rather, is influenced by some structural aspect of the protein that serves as a degradation signal, which is recognized and bound by an E3. The half-life of a protein is also influenced by the amino (N)-terminal residue,  the so-called N-end rule, and ranges from minutes to hours.

Destabilizing N-terminal amino acids include arginine and post-translationally modified amino acids such as acetylated alanine. In contrast, serine is a stabilizing amino acid. Additionally, proteins rich in sequences containing proline, glutamate, serine, and threonine  (called PEST sequences after the one-letter designations for these amino acids) are rapidly ubiquitinated and degraded and, therefore,  have short half-lives.