Molecular Cell Biology
Protein Folding, Modification and Degradation
In 1958, when the first x-ray crystallographic structural results of myoglobin, a water-soluble globular protein, were obtained, scientists were anticipating to see a structure as predictable and symmetrical as that of a double-stranded DNA molecule (Tooze et. al, 1991). Instead, they were met with a complex, asymmetric image. While at the time this was unexpected, today it is understood as a requirement for proteins. Since proteins must recognize many different types of molecules through detailed three-dimensional interactions, it seems sensible to have a diverse and irregular structure to accommodate a wide variety of functions. Nevertheless, there are still regular features of protein structures that are common to all proteins, the most important of which is secondary structure. While primary structure dictates the linear sequence of amino acids and the location of covalent linkages between the amino acids, secondary structure determines the folding and coiling of the protein through hydrogen bonding. For instance, in myoglobin, the interior is hydrophobic, maintained by the in-folding of hydrophobic side chains of the amino acids, while the hydrophilic part of the polypeptide face outwards forming hydrogen bonds with water molecules, thus remaining soluble. Such arrangements are highly complex and are primarily achieved through formation of a-helices, triple helices and b-pleated sheets. In the myoglobin example, the structure is maintained by 8 amphipathic a-helices, the hydrophobic sides of which face the interior while the hydrophilic sides face outwards (Lark et. al, 1989).
Notwithstanding, secondary structures are only folding intermediates that initiate and facilitate the formation of super-secondary, tertiary and quaternary structures (Wright et. al, 1998). A fitting example is that of a well-defined helical hairpin (a motif comprised of a helix-turn-helix) between helices G and H of myoglobin, which makes it a folding initiation site (Waltho et. al, 1993 and Matheson et. al, 1978). Another example is that of Src protein kinase having 4 protein domains; SH2 domain, SH3 domain and two kinase domains. These are substructures, produced by parts of the polypeptide chain that can fold independently into compact, stable structures (Alberts et. al, 2002). The central core of a domain can be constructed from a-helices, from b-sheets, or from combinations of these two fundamental folding elements e.g. the SH2 domain is comprised of (Xu et. al, 1997) a central antiparallel b-sheet with a-helices on either side (Tian et. al, 2011) whereas the SH3 domain is made up of five or six beta strands arranged as two tightly packed antiparallel b-sheets (Xue et. al, 2011). Each different combination is known as a protein fold. The smallest protein molecules contain only a single domain, whereas larger proteins can contain as many as several dozen domains, each having a distinct function. As the amino acids in a polypeptide further interact with each other through disulfide bonds, salt bridges, hydrogen bonds and hydrophobic interactions, the tertiary structure of the protein is assumed (Nishikawa et. al, 1972). Such interactions allow the individual secondary structures to form either globular or fibrous domains. The three-dimensional protein structure of insulin forms three disulfide bridges between thiol (-SH) groups on cysteine residues (Qiao et. al, 2001). If a protein contains more than one polypeptide chain, the interaction between each polypeptide monomer gives rise to the quaternary structure of the protein. For instance, a molecule of hemoglobin assumes quaternary structure when its 2 alpha-subunits and 2 beta-subunits interact with each other giving it a globular shape.
Protein folding in vivo is facilitated by the molecular chaperones and chaperonins. BIP is an example of a molecular chaperone in the endoplasmic reticulum. Another example is that of an integral protein in the endoplasmic reticulum called calnexin (CNX), which retains unfolded or unassembled N-linked glycoproteins in the endoplasmic reticulum, ensuring that only properly folded and assembled proteins progress to the secretory pathway. Improper folding may give rise to slowly developing diseases such as Alzheimer’s disease. Nearly every protein undergoes modification following synthesis, which may alter its activity, life span or its location in the cell . For instance, in the case of insulin, there is cleavage of the polypeptide into three parts, only two of which join to give the active form comprising 51 amino acids. Another such process taking place in membrane proteins is prenylation (Shafer et. Al 1992), which is the addition of a hydrophobic molecule (3-methyl-2-but-2-en-1-yl) to the protein, which has been suggested to facilitate the attachment of the protein to the membrane. This is seen to take place in the modification of the enzyme farnesyltransferase, when the cysteine amino acid on the C-terminus of the enzyme is prenylated, making the enzyme active in cellular signaling (Kohl et. Al, 1993).
Equally important to protein synthesis, folding and modification is the process of protein degradation if the cell is to digest the protein and utilize the products or convert it to an inactive form etc. There are both extracellular and intracellular pathways to achieve this. In the extracellular activity of carboxypeptidases, the peptide bond of an amino acid residue is cleaved at the carboxy-terminal end. In ubiquitin-mediated proteolytic pathways, ubiquitin is tagged onto the internal lysine side chain of the protein, which is then targeted by proteosomes (Izzi and Attisano, 2004) and subsequently degraded.
In conclusion, the myriad of processes taking place in the cell are successfully performed due to the fact that the structure of the proteins is highly diverse. Folding and modification further add to this diversity allowing a limited number of amino acids to combine into thousands of proteins with distinct functions, thereby rendering these processes indispensable to the cell.
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