User:Chase Haven/Sandbox 1
From Proteopedia
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=====Indiscernible Regions===== | =====Indiscernible Regions===== | ||
| - | The elusive alpha-C termini regions were unable to be resolved. This was actually expected, as these regions are thought to be intrinsically disordered. A study in 2006 used the PONDR VL-XT algorithm (Predictor Of Natural Disordered Regions)and found that of the 866 residues of fibrinogen, 397 or 46% were predicted to be disordered. The longest stretch being 129 consecutive disordered residues. Large portions of these figures correspond with the alpha-C terminal regions. Interestingly, these regions have been shown to be involved in binding numerous proteins including those involved in coagulation (FXIII, other fibrin molecules) fibrinolysis (plasminogen, tissue-plasminogen activator), and others. The intrinsic disorder could be beneficial for fibrinogen by allowing a broad binding diversity, while also | + | The elusive alpha-C termini regions were unable to be resolved. This was actually expected, as these regions are thought to be intrinsically disordered. A study in 2006 used the PONDR VL-XT algorithm (Predictor Of Natural Disordered Regions)and found that of the 866 residues of fibrinogen, 397 or 46% were predicted to be disordered. The longest stretch being 129 consecutive disordered residues. Large portions of these figures correspond with the alpha-C terminal regions. Interestingly, these regions have been shown to be involved in binding numerous proteins including those involved in coagulation (FXIII, other fibrin molecules) fibrinolysis (plasminogen, tissue-plasminogen activator), and others. The intrinsic disorder could be beneficial for fibrinogen by allowing a broad binding diversity, while also stabilizing the protein in a variety of trauma-induced scenarios such as hypothermia and acidosis. The results of this algorithm also coincide with, up until the past decade, the distinct lack of available structural data concerning the different domains of fibrinogen, and likely contribute to the difficulty of crystalization attempts. |
| - | + | As it pertains to this crystal structure, it is also possible that these portions of the molecule were proteolytically removed sometime during purification, as has been observed in the literature. N-terminal segments of the alpha and B chains in the central globular region were also unresolved, but are known to be mobile. | |
| - | It is also possible that these portions of the molecule were proteolytically removed sometime during purification, as has been observed in the literature. N-terminal segments of the alpha and B chains in the central globular region were also unresolved, but are known to be mobile. | ||
=====Cation-Pi Interactions===== | =====Cation-Pi Interactions===== | ||
Revision as of 19:33, 22 April 2014
Contents |
Fibrinogen
|
Biological Role
The process of haemostasis is crucial to stemming blood loss following vascular injury. It involves a complex balance of pro- and anti-coagulant activities by a multitude of enzymes and cofactors that ultimately lead to a fibrin clot followed by attenuation of the coagulation response to restore normal blood flow. In short, the serine protease thrombin can be thought of as the star player: thrombin cleaves fibrinogen to fibrin (forming the clot) and also activates a trans-glutaminase (Factor XIII) which will create cross-links in the clot to enhance it's tensile strength. Furthermore, upon the formation of the clot thrombin also activates protein C as part of a negative feedback loop, which in turn degrades various cofactors and ultimately shuts down the coagulation response.[1][2]
Fibrinogen, the chief proteinaceous component of a clot, is a multidomain glycoprotein consisting of three pairs of peptide chains termed alpha, beta and gamma. It circulates in blood at concentrations generally ranging from 2 to 3 mg/mL. Vascular injury exposes tissue factor-bearing subendothelial cells to flowing blood, triggering a coagulation response in order to generate thrombin. This serine protease has a unique specificity for cleavage sites on fibrinogen alpha and beta chains, and when cleaved fibrinopeptides A and B are released. This results in the exposure of polymerization sites known as "knobs", which are complementary to "holes" on other fibrinogen molecules. This interaction is non-covalent but strong enough to support the spontaneous growth of fibrin fibers and ultimately a fibrin clot network capable of stemming blood loss in vivo.[1][2]
Fibrin clots are highly heterogeneous; numerous factors play a role in determining fiber diameter, types of branch points, and number of branching fibers per unit area. Some of these factors are well understood, such as the effects of higher thrombin or fibrinogen concentrations. Others are less understood, such as calcium and metal ion levels, circulating lipid content, coagulation or fibrinolytic protein levels, some studies have even shown smoking and diabetes may change the structural layout or integrity of fibrin clots.[1][2]
Basic Crystalographic Features
The heterogeneity of fibrin clots make them virtually impossible to crystallize, and fibrinogen itself has intrinsic properties that enabled it to elude crystalization efforts until 2009. Some of the interesting aspects of this crystal are discussed below.
Coiled-Coil Domains
The coiled-coil domains separate the two outer D domains from the central globular E domain. These coils are mostly helical, with a notable exception being the “out-loop” in gamma chains residues 70-76. Certain areas also have runs of polar amino acids, coinciding with a pronounced bending at this location. Three notable runs include Alpha: 99-107, Beta: 126-131, and Gamma: 69-77. These non-helical regions share structural similarity and conservation with that of previously-crystalized chicken fibrinogen, even though they have strikingly different amino acid sequences.
It has been proposed that the coiled coils are the source of (at least in part) fibrinogen's inherent "floppiness". The non-helical stretches may act as flexible hinges interspersed with the more rigid helical components. This also fits with data regarding the compressible and stretchable nature of fibrin gels. Also fitting to this "hinge hypothesis", are two known mutations that cause defective fibrin formation occur in the polar (nonhelical) regions.
Indiscernible Regions
The elusive alpha-C termini regions were unable to be resolved. This was actually expected, as these regions are thought to be intrinsically disordered. A study in 2006 used the PONDR VL-XT algorithm (Predictor Of Natural Disordered Regions)and found that of the 866 residues of fibrinogen, 397 or 46% were predicted to be disordered. The longest stretch being 129 consecutive disordered residues. Large portions of these figures correspond with the alpha-C terminal regions. Interestingly, these regions have been shown to be involved in binding numerous proteins including those involved in coagulation (FXIII, other fibrin molecules) fibrinolysis (plasminogen, tissue-plasminogen activator), and others. The intrinsic disorder could be beneficial for fibrinogen by allowing a broad binding diversity, while also stabilizing the protein in a variety of trauma-induced scenarios such as hypothermia and acidosis. The results of this algorithm also coincide with, up until the past decade, the distinct lack of available structural data concerning the different domains of fibrinogen, and likely contribute to the difficulty of crystalization attempts.
As it pertains to this crystal structure, it is also possible that these portions of the molecule were proteolytically removed sometime during purification, as has been observed in the literature. N-terminal segments of the alpha and B chains in the central globular region were also unresolved, but are known to be mobile.
Cation-Pi Interactions
Subsection 4
The of Rad51 has been shown to bind DNA, [1][3][4] and is thought to have significant disordered character.[4] It contains a conserved glycine residue at position 103, although this is not shared by the Drosophila melanogaster Rad51 protein. Mutation of this residue to glutamate results in a greatly reduced ability to bind both ssDNA as well as dsDNA. This defect then leads to a significant reduction in ATPase activity. G103 lies at the surface of the N-terminal domain facing the core ATPase site, yet crystal structures show that G103 is removed from the NTP binding site, so there is no direct interaction that could explain the loss of ATPase activity.[5]
Evolutionary Conservation
References
- ↑ Cite error: Invalid
<ref>tag; no text was provided for refs namedLiu - ↑ 2.0 2.1 2.2 Chi P, Van Komen S, Sehorn MG, Sigurdsson S, Sung P. Roles of ATP binding and ATP hydrolysis in human Rad51 recombinase function. DNA Repair (Amst). 2006 Mar 7;5(3):381-91. Epub 2006 Jan 4. PMID:16388992 doi:10.1016/j.dnarep.2005.11.005
- ↑ Cite error: Invalid
<ref>tag; no text was provided for refs namedChen - ↑ 4.0 4.1 Aihara H, Ito Y, Kurumizaka H, Yokoyama S, Shibata T. The N-terminal domain of the human Rad51 protein binds DNA: structure and a DNA binding surface as revealed by NMR. J Mol Biol. 1999 Jul 9;290(2):495-504. PMID:10390347 doi:10.1006/jmbi.1999.2904
- ↑ Cite error: Invalid
<ref>tag; no text was provided for refs namedConway
