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How the Linker Connecting the Two Kringles Influences Activation and Conformational Plasticity of Prothrombin.

Pozzi N, Chen Z, Di Cera E
J Biol Chem 291 6071-82 (2016)
Cited: 22 times
EuropePMC logo PMID: 26763231

Abstract

A flexible linker (Lnk2) composed of 26 amino acids connects kringle-1 to kringle-2 in the coagulation factor prothrombin. Recent studies point to Lnk2 as a key determinant of the structure and function of this zymogen. Using a combination of mutagenesis, structural biology, and single molecule spectroscopy, we show how Lnk2 influences activation and conformational plasticity of prothrombin. Scrambling the sequence of Lnk2 is inconsequential on activation, and so is extension by as many as 22 residues. On the other hand, below a critical length of 15 residues, the rate of prothrombin activation increases (10-fold) in the absence of cofactor Va and decreases (3-fold) in the presence of cofactor. Furthermore, activation by prothrombinase takes place without preference along the prethrombin-2 (cleavage at Arg(271) first) or meizothrombin (cleavage at Arg(320) first) pathways. Notably, these transitions in the rate and pathway of activation require the presence of phospholipids, pointing to an important physiological role for Lnk2 when prothrombin is anchored to the membrane. Two new crystal structures of prothrombin lacking 22 (ProTΔ146-167) or 14 (ProTΔ154-167) residues of Lnk2 document striking conformational rearrangements of domains located across this linker. FRET measurements of freely diffusing single molecules prove that these structural transitions are genuine properties of the zymogen in solution. These findings support a molecular model of prothrombin activation where Lnk2 presents the sites of cleavage at Arg(271) and Arg(320) to factor Xa in different orientations by pivoting the C-terminal kringle-2/protease domain pair on the N-terminal Gla domain/kringle-1 pair anchored to the membrane.

Reviews - 5edm mentioned but not cited (1)

  1. Structure of Coagulation Factor II: Molecular Mechanism of Thrombin Generation and Development of Next-Generation Anticoagulants. Chinnaraj M, Planer W, Pozzi N. Front Med (Lausanne) 5 281 (2018)

Articles - 5edm mentioned but not cited (10)

  1. Anti-prothrombin autoantibodies enriched after infection with SARS-CoV-2 and influenced by strength of antibody response against SARS-CoV-2 proteins. Emmenegger M, Kumar SS, Emmenegger V, Malinauskas T, Buettner T, Rose L, Schierack P, Sprinzl MF, Sommer CJ, Lackner KJ, Aguzzi A, Roggenbuck D, Frauenknecht KBM. PLoS Pathog 17 e1010118 (2021)
  2. How the Linker Connecting the Two Kringles Influences Activation and Conformational Plasticity of Prothrombin. Pozzi N, Chen Z, Di Cera E. J Biol Chem 291 6071-6082 (2016)
  3. Non-canonical proteolytic activation of human prothrombin by subtilisin from Bacillus subtilis may shift the procoagulant-anticoagulant equilibrium toward thrombosis. Pontarollo G, Acquasaliente L, Peterle D, Frasson R, Artusi I, De Filippis V. J Biol Chem 292 15161-15179 (2017)
  4. Structure of prothrombin in the closed form reveals new details on the mechanism of activation. Chinnaraj M, Chen Z, Pelc LA, Grese Z, Bystranowska D, Di Cera E, Pozzi N. Sci Rep 8 2945 (2018)
  5. Structural Architecture of Prothrombin in Solution Revealed by Single Molecule Spectroscopy. Pozzi N, Bystranowska D, Zuo X, Di Cera E. J Biol Chem 291 18107-18116 (2016)
  6. Predicting protein-membrane interfaces of peripheral membrane proteins using ensemble machine learning. Chatzigoulas A, Cournia Z. Brief Bioinform 23 bbab518 (2022)
  7. Discovery and characterization of 2 novel subpopulations of aPS/PT antibodies in patients at high risk of thrombosis. Chinnaraj M, Planer W, Pengo V, Pozzi N. Blood Adv 3 1738-1749 (2019)
  8. Probing prothrombin structure by limited proteolysis. Acquasaliente L, Pelc LA, Di Cera E. Sci Rep 9 6125 (2019)
  9. Cryo-EM structure of the prothrombin-prothrombinase complex. Ruben EA, Summers B, Rau MJ, Fitzpatrick JAJ, Di Cera E. Blood 139 3463-3473 (2022)
  10. The Fragment 1 Region of Prothrombin Facilitates the Favored Binding of Fragment 12 to Zymogen and Enforces Zymogen-like Character in the Proteinase. Bradford HN, Krishnaswamy S. J Biol Chem 291 11114-11123 (2016)


Reviews citing this publication (3)

  1. Plasminogen Activator Inhibitor Type-1 as a Regulator of Fibrosis. Rabieian R, Boshtam M, Zareei M, Kouhpayeh S, Masoudifar A, Mirzaei H. J Cell Biochem 119 17-27 (2018)
  2. Exosite Binding in Thrombin: A Global Structural/Dynamic Overview of Complexes with Aptamers and Other Ligands. Troisi R, Balasco N, Autiero I, Vitagliano L, Sica F. Int J Mol Sci 22 10803 (2021)
  3. Cryo-EM structures of coagulation factors. Di Cera E, Mohammed BM, Pelc LA, Stojanovski BM. Res Pract Thromb Haemost 6 e12830 (2022)

Articles citing this publication (8)

  1. Interplay between conformational selection and zymogen activation. Chakraborty P, Acquasaliente L, Pelc LA, Di Cera E. Sci Rep 8 4080 (2018)
  2. Enhancing the anticoagulant profile of meizothrombin. Stojanovski BM, Pelc LA, Zuo X, Pozzi N, Cera ED. Biomol Concepts 9 169-175 (2018)
  3. Residues W215, E217 and E192 control the allosteric E*-E equilibrium of thrombin. Pelc LA, Koester SK, Chen Z, Gistover NE, Di Cera E. Sci Rep 9 12304 (2019)
  4. Zymogen and activated protein C have similar structural architecture. Stojanovski BM, Pelc LA, Zuo X, Di Cera E. J Biol Chem 295 15236-15244 (2020)
  5. Role of sequence and position of the cleavage sites in prothrombin activation. Stojanovski BM, Di Cera E. J Biol Chem 297 100955 (2021)
  6. Dual effect of histone H4 on prothrombin activation. Pozzi N, Di Cera E. J Thromb Haemost 14 1814-1818 (2016)
  7. Fruit Bagasse Phytochemicals from Malpighia Emarginata Rich in Enzymatic Inhibitor with Modulatory Action on Hemostatic Processes. Marques TR, Cesar PHS, Braga MA, Marcussi S, Corrêa AD. J Food Sci 83 2840-2849 (2018)
  8. Comparative sequence analysis of vitamin K-dependent coagulation factors. Stojanovski BM, Di Cera E. J Thromb Haemost 20 2837-2849 (2022)


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