Disulfides between protein subunits

Figure 18-8 Stereoscopic ribbon diagrams of the chicken bc1 complex (A) The native dimer. The molecular twofold axis runs vertically between the two monomers. Quinones, phospholipids, and detergent molecules are not shown for clarity. The presumed membrane bilayer is represented by a gray band. (B) Isolated close-up view of the two conformations of the Rieske protein (top and long helix at right) in contact with cytochrome b (below), with associated heme groups and bound inhibitors, stigmatellin, and antimycin. The isolated heme of cytochrome c, (left, above) is also shown. (C) Structure of the intermembrane (external surface) domains of the chicken bcx complex. This is viewed from within the membrane, with the transmembrane helices truncated at roughly the membrane surface. Ball-and-stick models represent the heme group of cytochrome cy the Rieske iron-sulfur cluster, and the disulfide cysteines of subunit 8. SU, subunit cyt, cytochrome. From Zhang et al.105...

SDS disrupts noncovalent interactions between subunits of a protein, so if a protein has two subunits, two bands will appear. In the absence of SDS, only one band will appear. This reagent and mercaptoethanol reduce protein subunits that are disulfide bonded. This property of SDS may be responsible for protein denaturation. It should be noted that SDS also permeabilizes cells for antibody access to intracellular epitopes. 

Sodium Dodecyl Sulfate-Polyacrylamide Gel Electrophoresis (SDS-PAGE). Sodium dodecyl sulfate (SDS) gel electrophoresis systems are used to determine the number and size of protein chains or protein subunit chains in a protein preparation. Initially, the protein preparation is treated with an excess of soluble thiol (usually 2-mercaptoethanol) and SDS. Under these conditions, the thiol reduces all disulfide bonds (-S-S-) present within and/or between peptide units, while the SDS (an ionic or... 

Interactions of secondary structures within a single polypeptide chain generate tertiary structuresthe complex three-dimensional shapes of proteins. Interactions between separate polypeptide chains in the same protein generate a quaternary structure, which mediates interactions between the subunits. Tertiary and quaternary structures may be stabilized by disulfide bridges between cysteine residues, forming cystines. 

Finally, the structure of the whole protein including the disulfide bridges between different subunits can be determined. 

Fibronectin is an adhesion protein like laminin, vitronectin, and von Wille-brand factor, which are synthesized by the cells themselves to build up the ECM. The glycoprotein fibronectin with a molecular weight between 220,000 and 250,000 consists of two similar subunits, which are connected close to their C-terminus by disulfide bridges. The subunits are composed of functional domains [121]. The cell binding domain with the characteristic sequence Gly-Arg-Gly-Asp-Ser (GRGDS) is of special interest [122]. Models of the subunit of the fibronectin molecule and its cell binding domain are presented in Fig. 21. 

Figure 11.6. Schematic diagram showing the assembly of IL-12 protein for antibody-based drug delivery, (a) The mature sequences of the p35 subunit of IL-12 are fused to the C-terminus of the heavy chain of a tumour-specific antibody and co-expressed with the antibody light chain and the p40 subunit of IL-12. Formation of the final immunocytokine requires the creation of disulfide bridges between the antibody chains and interactions of p35 and p40 subunits of IL-12 [119]. (b) Alternatively the IgG heavy chain and both subunits of IL-12 can be linked via flexible linkers allowing for equimolar assembly of IL-12 [120].

Larger proteins often contain more than one polypeptide chain. These multi-subunit proteins have a more complex shape, but are still formed from the same forces that twist and fold the local polypeptide. The unique three-dimensional interaction between different polypeptides in multi-subunit proteins is called the quaternary structure. Subunits may be held together by noncovalent contacts, such as hydrophobic or ionic interactions, or by covalent disulfide bonds formed from the cysteine residue of one polypeptide chain being cross-linked to a cysteine sulfhydryl of another chain (Fig. 15). 

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