Homodimer, noncovalent

Neurotrophins (NGF brain-derived neurotrophic factor, BDNF neurotrophin-3, NT-3 NT-4 NT-6) are important regulators of neural survival, development, function, and plasticity of the vertebrate nervous system [1]. Neurotrophins generally function as noncovalently associated homodimers. They activate two different classes of receptors, through which signaling pathways can be activated, including those mediated by Ras and members of the cdc42/rac/rho G protein families, MAP kinase, PI-3 kinase, and Jun kinase cascades. 

IL-20. IL-20 is a novel member of the expanding family of IL- 10-like cytokines, which includes the other novel cytokines IL-19 and IL-22 (B11, XI). It is presumed that the bioactive form of IL-20 is a noncovalently linked homodimer, similar to IL-10. 

PDI is a noncovalent homodimer with a molecular weight of 57 kDa for each subunit. The protein is generally involved in the formation, reduction and rearrangement of disulfide bridges and plays an important role in platelet function. PDI is localized at the extracellular side of the plasma membrane and is secreted after platelet activation (Chen etal, 1992). In platelets, PDI catalyzes the formation of disulfide-bridged complexes of thrombospondin 1 and thrombin-antithrombin III and is thus involved in hemostasis and wound healing (Milev and Essex, 1999). 

We have been studying the E.coli expression of human neurotrophin-3 (NT-3), a member of the nerve growth factor (NGF) family of neurotrophic factors. The protein is believed to have value in the treatment of certain neurodegenerative diseases (6). Mature NT-3 is a polypeptide of 119 amino acid residues and under physiological conditions, the protein functions as a tightly associated, noncovalently liiiked homodimer. Expression of NT-3 using UAA or UAG as the... 

The bacterial, iron-containing cd nitrite reductases constitute another family of enzymes catalyzing the one-electron reduction of nitrite to nitric oxide (74). These enzymes are homodimers of 60-kDa subunits, each containing one heme-c and one heme-rii. Extensive studies have established heme-c as the electron entry site, whereas heme-dj is the catalytic center where nitrite is reduced (95). Three-dimensional structures of two different cytochromes cd have been determined in oxidized and reduced states P. pantotrophus, Pp-NiR (96, 97) and P. aeruginosa, Pa-NiR (98, 99). In both enzymes, heme-c is covalently linked to the A-terminal a-helical domain and heme-di is bound noncovalently to the C-terminal B-propeller domain (Fig. 17). Intramolecular ET between c and di hemes is an essential step in the catalytic cycle this reaction has been studied by several groups using different methods (95, 100-106). The rate constants for Pa-NiR are on the order of 1 s (95, 100-102, 104), while intramolecular ET in Pp-NiR is significantly faster (rate constant of... 

Two types of 15-kD subunits are present, one type beginning with N-terminal Ala and the other with N-terminal Leu. Pairs of these small 15-kD subunits are linked by covalent disulfide bonds that are broken only by the mercaptoethanol. The disulfide-linked subunits comprise the 30-kD species. (There is insufficient information to discern whether the 30-kD species consists of two different homodimers in which identical subunits are linked by disulfide bonds, or a unique heterodimer in which one Ala-initiated subunit is linked precisely to one Leu-initiated subunit.) Finally, two of the 30-kD species associate noncovalently to form the 60-kD particle that contains two copies each of two different 15-kD subunits. The 60-kD particle constitutes the native protein that is observed by molecular exclusion chromatography. (Urea disrupts the noncovalent subunit association, without breaking the disulfide bonds.)... 

Examples from other groups include gas-phase thermal dissociation experiments, implemented with blackbody infrared radiative dissociation (BIRD) and FT-ICR MS on a series of protein-carbohydrate complexes, and the detection of fusion peptide-phospholipid noncovalent interactions using nano-ESI FTICR-MS. An interesting example of protein-DNA interaction smdied by ESI-MS is the trp repressor-DNA operator complex. Escherichia coli trp apo-repressor (TrpR), a homodimer, is a DNA-binding protein that binds to two molecules of co-repressor L-tryptophan to form a holorepressor complex at higher salt concentrations. The mass spectrum of noncovalent... 

Such ESl-MS measurements of the noncovalent complexes can be helpful to confirm protein function by defining the quaternary structure. The putative Spodoptera frugiperda glutathione S-transferase described in Figures 18.2-18.4was measured by nanoelectrospray MS using near-neutral pH solvents to help retain their native structures. The unknown protein shows a protein dimer as the most stable complex, consistent with the function of GST proteins from other species. From the three distinct protein monomers with molecular masses approximately 23 kDa measured by ESl-MS at pH 2.5, at least six hetero- and homodimers of 46 kDa are measured from a pH 7.5 solution (Figure 18.10). Nearly all GST... 

In the structure of pyruvate dehydrogenase complex, the E2 component forms the central core of the complex. PDHc from Gram-negative bacteria possesses an octahedral E2 core while the Gram-positive and eukaryotic PDHc are based on an icosahedral core [22]. The El and E3 components are attached noncovalently to the E2 core. The El component occurs in two forms depending on the symmetry of the complex. In octahedral complexes, El component is an (a,) homodimer with a subunit mass of approximately 100 kDa, while in icosahedral complexes, it exists as an (a2P2) heterotetramer with subunits of approximately 41 kDa and 36 kDa [23,24]. 

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