Allosteric Disulfide Bonds in Thrombosis and Thrombolysis
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Journal of Thrombosis and Haemostasis, 4: 2533–2541 REVIEW ARTICLE Allosteric disulfide bonds in thrombosis and thrombolysis V . M . C H E N * and P . J . H O G G * *Centre for Vascular Research, University of New South Wales, Sydney; and Children’s Cancer Institute Australia for Medical Research, Sydney, Australia To cite this article: Chen VM, Hogg PJ. Allosteric disulfide bonds in thrombosis and thrombolysis. J Thromb Haemost 2006; 4: 2533–41. most frequently acquired amino acid in eight of the 15 taxa Summary. Allosteric disulfide bonds control protein function studied. Other amino acids that have accrued are Met, His, Ser by mediating conformational change when they undergo and Phe, whereas Pro, Ala, Glu and Gly have been lost. reduction or oxidation. The known allosteric disulfide bonds Considering that disulfide bonds follow addition of Cys, this are characterized by a particular bond geometry, the )RHSta- analysis indicates that acquisition of these bonds is a relatively ple. A number of thrombosis and thrombolysis proteins contain recent evolutionary event. one or more disulfide bonds of this type. Tissue factor (TF) was the first hemostasis protein shown to be controlled by an Types and classification of disulfide bonds allosteric disulfide bond, the Cys186–Cys209 bond in the membrane-proximal fibronectin type III domain. TF exists in There are two general types of disulfide bond; structural and three forms on the cell surface: a cryptic form that is inert, a functional. The structural bonds, which are the majority, are coagulant form that rapidly binds factor VIIa to initiate involved in the folding of a protein and stabilize the tertiary coagulation, and a signaling form that binds FVIIa and cleaves structure. It is thought that they assist folding by decreasing the protease-activated receptor 2, which functions in inflamma- entropy of the unfolded form [4]. The functional disulfide tion, tumor progression and angiogenesis. Reduction and bonds are the catalytic and allosteric bonds. These disulfide oxidation of the Cys186–Cys209 disulfide bond is central to the bonds regulate protein function. transition between the three forms of TF. The redox state of the The catalytic bonds are the better characterized. Examples of bond appears to be controlled by protein disulfide isomerase these are the disulfide bonds in the active sites of oxidoreduc- and NO. Plasmin(ogen), vitronectin, glycoprotein 1ba,integrin tases, such as the thioredoxin family of proteins [5,6]. These b3 and thrombomodulin also contain )RHStaple disulfides, disulfide bonds cycle between reduced and oxidized configu- and there is circumstantial evidence that the function of these rations in coordination with a dithiol or disulfide in a protein proteins may involve cleavage/formation of these disulfide substrate, resulting in reduction, formation or isomerization of bonds. a disulfide in the substrate. The allosteric bonds have only recently been identified [7]. These bonds control protein Keywords: glycoprotein 1ba,integrinb3, plasminogen, thrombo- function by triggering a conformational change when they modulin, tissue factor, vitronectin. break and/or form and are the subject of this review. A disulfide bond can be described in terms of the five Introduction dihedral or v angles that make up the bond. A dihedral angle is the angle of rotation about a certain bond, and is defined using Protein disulfide bonds are covalent links between two Cys four atoms [8]. v1 is the dihedral angle about the Ca–Cb bond, residues in the polypeptide chain. It appears that all life-forms v2 about the Cb–Sc bond, v3 about the Sc–Sc¢ bond, v2¢ about make this bond. Mammalian cells, for instance, make about the Sc¢–Cb¢ bond and v1¢ about the Cb¢–Ca¢ bond. These five 3000 different proteins that contain disulfide bonds. angles can be used to estimate the potential energy of a disulfide Addition of disulfide bonds to proteins is a major mechan- bond, which is called the dihedral strain energy (DSE) [8–10]. ism by which proteins have evolved and are still evolving [1–3]. The DSE is a semiquantitative measure, as it does not factor in Analysis of the trend in amino gain and loss in protein bond lengths and van der Waals contacts, for example, but has evolution has shown that Cys residues have accrued in 15 taxa been shown experimentally to reflect the amount of strain in a representing all three domains of life [3]. Moreover, Cys was the disulfide bond [11–14]. There are three basic disulfide bond configurations, based on Correspondence: Philip J. Hogg, Centre for Vascular Research, the sign of the v2, v3 and v2¢ angles; the spirals, hooks or staples University of New South Wales, Sydney NSW 2052, Australia. [15]. If the v3 angle is positive, then the bond is right-handed; if Tel.: +61 2 9385 1004; fax: +61 2 9385 1389; e-mail: it is negative, the bond is left-handed. When the v1 and v1¢ [email protected] angles are considered, there are 20 possible disulfide bond Ó 2006 International Society on Thrombosis and Haemostasis 2534 V. M. Chen and P. J. Hogg configurations. All 20 disulfide bond geometries were identified likely mediator [20], although protein disulfide isomerase (PDI) in a dataset of 6874 unique disulfide bonds in X-ray structures has also been implicated [24]. [8]. The main structural disulfide bond is the )LHSpiral. A In this review, we will discuss the presence of allosteric quarter of the disulfide bonds in the dataset were )LHSpirals, disulfide bonds in proteins involved in thrombosis and and this group also had the lowest DSE. A striking finding thrombolysis, and evidence for control of some of these from this analysis was that nearly all the catalytic disulfide proteins by these bonds (Table 1). We will focus initially on bonds were of the +/)RHHook configuration, whereas all the tissue factor (TF), which is the best characterized example to known allosteric disulfide bonds fell into the )RHStaple group. date. Other proteins that will be discussed are plasmin(ogen), Both of these bond groups have a high and narrow DSE vitronectin, glycoprotein 1ba,integrinb3 and thrombo- distribution, which is consistent with their functional role. modulin. A defining feature of the )RHStaple group is the short distance between the a carbon atoms of the disulfide bond. Tissue factor )RHStaple bonds have a mean Ca–Ca¢ distance of 4.3 A˚ , compared to a mean of 5.6 A˚ for all disulfide bonds [8]. This The blood coagulation process is initiated when the circulating short Ca–Ca¢ distance results from the fact that )RHStaples serine protease, factor (F) VIIa, forms a complex with its mostly link adjacent strands in the same antiparallel b-sheet cofactor, membrane-bound TF, to activate FVII, FX and FIX [7,16]. The strands need to pucker to accommodate the by limited proteolysis. Ternary TF–FVIIa–FXa or binary disulfide bond, and the associated deformation of the b-sheet TF–FVIIa complexes also signal through cleavage of protease- imparts a high torsional energy on the bond. A number of activated receptor 2 (PAR2) to control inflammation, tumor )RHStaple disulfide bonds have been shown to be involved in progression and angiogenesis. TF is a transmembrane glyco- the function of the protein in which they reside. The best protein with structural homology to the class II cytokine characterized are the )RHStaple disulfide bonds in the superfamily. It consists of two fibronectin type III domains, bacterial proteins botulinum neurotoxins [16,17], PapD [18] each with a disulfide bond. and DsbD [19], and the mammalian immune and human Analysis of 16 X-ray TF structures indicated that the immunodeficiency virus (HIV) receptor CD4 [20,21]. The viral N-terminal domain Cys49–Cys57 disulfide bond exists in one envelope glycoprotein, HIV gp120, also appears to be regulated of two configurations: a )RHSpiral or a +/)RHSpiral by cleavage of one or more of its four )RHStaple disulfide (Table 2, Fig. 1). Both configurations are subgroups of the bonds [16,22,23]. Analysis of the )RHStaple disulfide bonds in main structural disulfide bond, the )LHSpiral [8]. This type of nuclear magnetic resonance (NMR) structures indicates that disulfide bond is typical of immunoglobulin-like domains, of theyoftenswitchtothe)LHStaple configuration, which has an which the fibronectin type III domain is an example [25]. The even higher mean torsional energy and shorter mean Ca–Ca¢ disulfide bond links across the two sheets of the b-sandwich distance than the )RHStaple bonds (B. Schmidt and P. J. structure. The C-terminal domain Cys186–Cys209 disulfide Hogg, unpubl. obs.). The )LHStaple bonds, therefore, should bond exists exclusively as a )RHStaple in the 16 structures [26]. also be considered as potential allosteric disulfide bonds. In This bond has a higher DSE and a considerably shorter Ca– fact, it is an open question whether the )RHStaple or Ca¢ distance than the N-terminal RHSpiral (Table 2), which is )LHStaple configuration is the functional form in some in accordance with the typical features of these bonds [8]. The proteins. Cys186–Cys209 disulfide bond straddles the F and G strands of CD4 is a good example of how protein function can be the antiparallel b-sheet at a non-H-bonded site. The disulfide controlled by an allosteric )RHStaple disulfide bond. CD4 is a bond constrains a b hairpin turn adjacent to the stalk region class II cytokine receptor that is expressed on T cells. It is a coreceptor for binding of MHCII to the T-cell receptor and is the primary receptor for HIV. The extracellular part of CD4 Table 1 RHStaple disulfide bonds in selected thrombosis and thrombo- consists of a concatenation of four immunoglobulin-like lysis proteins domains, and the first, second and fourth domains contain a Protein Region Disulfide bond(s)* single disulfide bond.