Proteins N-terminus

Figure 42 The structure of apoferritin. N = N-terminus protein helices E form hydrophobic channels (reproduced with permission from Adv. Inorg. Biochem., 1984, 5, 39, Elsevier, Amsterdam)...

N terminus (Section 27 7) The amino acid at the end of a pep tide or protein chain that has its a ammo group intact that IS the a ammo group is not part of a peptide bond... 

The methods involved in the production of proteins in microbes are those of gene expression. Several plasmids for expression of proteins having affinity tails at the C- or N-terminus of the protein have been developed. These tails are usefiil in the isolation of recombinant proteins. Most of these vectors are commercially available along with the reagents that are necessary for protein purification. A majority of recombinant proteins that have been attempted have been produced in E. Coli (1). In most cases these recombinant proteins formed aggregates resulting in the formation of inclusion bodies. These inclusion bodies must be denatured and refolded to obtain active protein, and the affinity tails are usefiil in the purification of the protein. Some of the methods described herein involve identification of functional domains in proteins (see also Protein engineering). 

Figure 2.2 The a helix is one of the major elements of secondary structure in proteins. Main-chain N and O atoms ate hydrogen-bonded to each other within a helices, (a) Idealized diagram of the path of the main chain in an a helix. Alpha helices are frequently illustrated in this way. There are 3.6 residues per turn in an a helix, which corresponds to 5.4 A (1.5 A pet residue), (b) The same as (a) but with approximate positions for main-chain atoms and hydrogen bonds Included. The arrow denotes the direction from the N-terminus to the C-termlnus.

The fundamental unit of tertiary structure is the domain. A domain is defined as a polypeptide chain or a part of a polypeptide chain that can fold independently into a stable tertiary structure. Domains are also units of function. Often, the different domains of a protein are associated with different functions. For example, in the lambda repressor protein, discussed in Chapter 8, one domain at the N-terminus of the polypeptide chain binds DNA, while a second domain at the C-terminus contains a site necessary for the dimerization of two polypeptide chains to form the dimeric repressor molecule. 

An exopeptidase that sequentially releases an amino acid from the N-terminus of a protein or peptide. Examples include cystinyl aminopeptidase (MEROPS M01.011), which removes a terminal cysteine from the biologically important peptides oxytocin and vasopressin, and methionyl aminopeptidase (M24.001), which removes the initiating methionine from cytosolic... 

The a subunits, for which two isoforms exist in mammals (al, a2), contain conventional protein serine/threonine kinase domains at the N-terminus, with a threonine residue in the activation loop (Thr-172) that must be phosphorylated by upstream kinases (see below) before the kinase is active. The kinase domain is followed by an autoinhibitory domain, whose effect is somehow relieved by interaction with the other subunits. The C-terminal domain of the a subunit is required for the formation of a complex with the C-terminal domain of the (3 subunit, which in turn mediates binding to the y subunit. The al and a2 catalytic subunit isoforms are widely distributed, although a2 is most abundant in muscle and may be absent in cells of the endothelial/hemopoietic lineage. 

Important members of this toxin family are Clostridium difficile toxins A and B, which are implicated in antibiotics-associated diarrhea and pseudomembranous colitis. The large clostridial cytotoxins are single-chain toxins with molecular masses of 250-308 kDa. The enzyme domain is located at the N terminus. The toxins are taken up from an acidic endosomal compartment. They glucosylate RhoA at Thr37 also, Rac and Cdc42 are substrates. Other members of this toxin family such as Clostridium sordellii lethal toxin possess a different substrate specificity and modify Rac but not Rho. In addition, Ras subfamily proteins (e.g., Ras, Ral, and Rap) are modified. As for C3, they are widely used as tools to study Rho functions [2] [4]. 

The presence of chromosomal translocations is a consistent feature of many leukemia s, lymphomas, and certain solid tumors. At the genetic level, these events can either deregulate an intact gene by disruption or removal and replacement of the adjacent controlling elements, or create a new fusion gene that express the N-terminus of one protein fused to the C-terminus of another protein. 

Threonine peptidases (and some cysteine and serine peptidases) have only one active site residue, which is the N-terminus of the mature protein. Such a peptidase is known as an N-terminal nucleophile hydrolase or Ntn-hydrolase. The amino group of the N-terminal residue performs the role of the general base. The catalytic subunits of the proteasome are examples of Ntn-hydrolases. 

Cleavage occur s at the scissile bond. Residues in the substrate towards the N-terminus are numbered PI, P2, P3, etc, whereas residues towards the C-terminus are numbered PI, P2, P3 etc. Cleavage occurs between PI and P1. For a peptidase with limited specificity, only the residue in PI or PI is important for specificity. A peptidase with an extended substrate binding site will have a preference for residues in other positions. For example cathepsin L prefers substrates with phenylalanine in P2 and arginine in PI. However, this is a preference only, and cathepsin L cleaves substrates after other amino acids. Caspase-3 has a preference for Asp in both P4 and PI, but it is unusual for substrate specificity to extend much further from the scissile bond. The peptidase with the most extended substrate specificity may be mitochondrial intermediate peptidase that removes an octopeptide targeting signal from the N-terminus of cytoplasmically synthesized proteins that are destined for import into the mitochondrial lumen. 

An enzyme in which the single catalytic residue is at the N-terminus of the protein. Many Ntn-hydrolases are synthesized as precursors and autoactivate the precursors are therefore peptidases, even if the mature enzyme has no further proteolytic activity. Three of the beta subunits of the proteasome are Ntn-hydrolases. 

Cell membrane spanning proteins contain a luminal/ extracellular domain, a transmembrane region and a cytosolic domain. In a type I transmembrane protein the N-terminus is the extracellular/luminal part of the protein, whereas the C-terminus comprises the cytosolic region of the membrane protein. 

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