BIOCHEMISTRY - Textbook - Ostapchenko L. I. - 2012

Chapter 6. AMINO ACID METABOLISM AND FUNCTIONS. PROTEIN BIOSYNTHESIS

6.10. Protein biosynthesis

6.10.9. Protein Folding and Processing

At the final Stages of the biosynthetic process, Polypeptides fold into their secondary and tertiary Conformations and undergo Processing. This process, which can occur both cotranslationally (simultaneously with protein Translation in the ribosome) and posttranslationally (after the completion of translation and the release of the protein from the ribosome), includes:

✵ folding of the peptide chain to form a unique tertiary or quaternary Structure;

✵ Modification of protein molecules;

✵ delivery of the protein to its site of future functioning.

Folding. At The final stage of Protein Synthesis, the polypeptide chain folds to acquire its native conformation through hydrogen, hydrophobic, Van der Waals, ionic, and covalent interactions. This process can be spontaneous, as proven by C. Anfinsen's experiments in the late 1950s.

For A number of Proteins, correct polypeptide chain folding is carried out under the control of specialized folding Enzymes (foldases) and chaperones, primarily heat Shock proteins (Hsp). By binding to the peptide molecule and activating ATP-dependent processes, chaperones ensure the proper folding of synthesized proteins, enable partial refolding, and participate in various Intracellular Transport processes.

Posttranslational modifications. Structural Proteins and Enzymes can be modified covalently and noncovalently at both the N- and C-termini of the molecule, as well as at amino acid side chains. These include: Acetylation of the N-terminal amino acid in the protein molecule; hydroxylation of the Proline residue during the transition from procollagen to Collagen; methylation of Lysine and Arginine residues using S-adenosyl-L-Methionine; attachment of oligosaccharide fragments to asparagine, Serine, and Threonine residues during glycoprotein Biosynthesis (glycosylation); amidation of aspartic and glutamic acid residues, associated with protein Variability during ontogenesis, particularly during Aging; carboxylation of glutamic acid residues, resulting in The formation of $\gamma$-carboxyglutamic acid residues in proteins, which are required, in particular, for Ca²⁺ binding; adenylylation and uridylylation of Tyrosine residues. Phosphorylation of Histones and non-histone Chromatin proteins is a posttranslational modification of major importance for regulating the METABOLIC ACTIVITY OF The Genome. Phosphorylation at

hydroxyl groups of serine, threonine, and tyrosine, as well as dephosphorylation, is believed to be involved in a wide range of metabolic processes. In addition, modification of amino acid residues can occur, such as The conversion of lysine and proline into hydroxylysine and hydroxyproline in collagen, the Iodination of tyrosine in thyroglobulin, and so on.

In Eukaryotic Cells, many proteins are synthesized as precursor molecules that require modification to acquire biological activity. Insulin, for example, is synthesized as proinsulin and is a single-chain molecule. The pathways of its modification involve proteolytic Cleavage and disulfide bond formation. After the removal of the polypeptide peptide segment by specific proteases, insulin is converted into a two-chain molecule with intra- and interchain disulfide bridges. Therefore, correctly formed Disulfide Bonds are crucial for maintaining the native conformation of molecules. A number of enzymes (trypsinogen, chymotrypsinogen) undergo partial proteolysis. Modifications of the N-terminus of synthesized Peptides occur through the action of deformylases and specific aminopeptidases. The attachment of a prosthetic group to form Conjugated Proteins and the assembly of protomers into an oligomeric protein are also stages of posttranslational modifications.

Protein splicing is an intramolecular autocatalytic process characteristic of archaea, Bacteria, and eukaryotes. During protein splicing, a specific internal fragment (intein) is excised, and the two remaining terminal fragments (exteins) are ligated to form a functional protein molecule. This process requires neither Energy Sources nor additional enzymes and Cofactors.

Protein targeting. As noted earlier, some of the proteins synthesized by a Introduction/5.html">Eukaryotic Cell are directed to their specific localization sites depending on their functional purpose. Lysosomal proteins, export proteins (secreted by The Cell), and plasma Membrane Proteins are synthesized on Endoplasmic reticulum (ER)-bound Ribosomes. Cytosolic proteins are synthesized on free ribosomes and remain at the site of synthesis, whereas protein targeting to Mitochondria, METABOLISM/14.html">Chloroplasts, and nuclei occurs via three distinct pathways. Elucidation of the molecular mechanisms has shown that these processes are mediated by signal sequences. This hypothesis was first proposed in 1970 by G. Blobel and D. Sabatini and later became known as the "Blobel signal hypothesis". Today, this hypothesis has been experimentally confirmed.

Using proteins synthesized on ER-bound ribosomes as an example, let us examine protein targeting. Signal sequences are fragments of polypeptide chains (15–30 amino acid residues) that are synthesized first from the N-terminus upon decoding signal codons located immediately after the initiation codons. In their central region, they contain hydrophobic amino acid residues, while their terminal sequences contain hydrophilic amino acid residues. This structure, provided there are at least 7 hydrophobic residues in a row, ensures the unobstructed penetration of signal peptides into the lipoprotein membrane at the signal peptide receptor site, followed by the translocation of the protein across The endoplasmic reticulum membrane or its anchorage within it. It has been established that protein translocation across Biological Membranes can occur cotranslationally or posttranslationally, and in both cases, the signal peptide plays a leading role, being cleaved upon completion of the process by signal peptidase.

Blobel's signal hypothesis stimulated research into the active Transport of Macromolecules: the existence of specialized proteins—porins—that facilitate macromolecule transport has been proven; membrane proteins that recognize signal peptides have been characterized; and signal-recognition particles (SRPs), which are ribonucleoprotein particles controlling the passage of the synthesized peptide across the membrane, have been identified.

Intracellular proteolysis. Proteins vary significantly in their turnover rate, namely Synthesis and degradation. An indicator of this is the protein half-life—the period during which 50% of these molecules are degraded. Active Proteolytic Enzymes are localized mainly in Lysosomes, which contain proteinases activated at acidic pH values—cathepsins (A, B, C, D, E). Neutral proteinases operating primarily in the Cytoplasm include calpains, which are Ca²⁺-dependent proteinases whose activity is associated with both structural and Metabolic Transformations in the cell. A special group of proteinases with an alkaline pH optimum is represented by kallikreins, whose activity is observed mainly in the Blood. The action of plasma kallikreins generates the peptide bradykinin, which regulates blood flow and cell membrane permeability. In recent years, The Role of ubiquitination as a determinant in Protein metabolism has been established. The attachment of ubiquitin (74 aa, molecular mass 8.5·10³) to the $\varepsilon$-amino group of a lysine residue in a protein destined for degradation targets it to the multisubunit proteolytic complex known as the proteasome. Protein molecule destruction is highly selective. The presence of leucine, lysine, or arginine at the N-terminus promotes protein degradation, whereas, for example, methionine, Alanine, and serine increase protein half-life. The presence of specific Amino acid sequences (proline (P), glutamic acid (E), serine (S), threonine (T)—PEST sequences) and oxidized amino acid residues in the protein molecule promotes the acceleration of its proteolytic degradation.