Characterization of the binding of a glycosylated serine protease from Euphorbia cf. lactea latex to human fibrinogen.
In this study, the binding of a glycosylated serine protease (EuP-82) with human fibrinogen was investigated by isothermal titration calorimetry (ITC). ITC analysis indicated that the binding of EuP-82 to fibrinogen in the conditions with or without the activator (Ca2+) was an exothermic reaction (dominant negative enthalpy), which tended to be driven by hydrogen bonding and van der Waals interactions. In contrast, the binding of fibrinogen-EuP-82 in the condition with the inhibitor (Zn2+) was an unfavorable endothermic reaction. EuP-82 could not inhibit the platelet activity in citrated whole blood via the ADP-receptor pathways (mainly, P2Y1 and P2Y12), but it could enhance the platelet aggregation. The ITC together with whole blood platelet aggregation suggested that EuP-82 provided multiple fibrinogen-binding sites that were not related to the arginine-glycine-aspartate (RGD) and the dodecapeptide sequences of fibrinogen. In addition, EuP-82 had neither thrombin-like activity nor anticoagulant activity. The SR-FTIR spectra revealed that EuP-82 was a glycoprotein. Deglycosylation of EuP-82 did not affect its proteolytic activity. Moreover, EuP-82 did not exhibit any toxicity to the living cells (NIH-3T3). This study supports that EuP-82 may be useful for wound-healing material through stabilizing the clot via the platelet induction for the first process.
School of Chemistry, Institute of Science, Suranaree University of Technology, Nakhon Ratchasima, Thailand.
Jaruwan
J
Siritapetawee
School of Chemistry, Institute of Science, Suranaree University of Technology, Nakhon Ratchasima, Thailand.
Chutima
C
Talabnin
National Center for Genetic Engineering and Biotechnology, National Science and Technology Development Agency, Thailand Science Park, Khlong Luang, Pathum Thani, Thailand.
Jarunee
J
Vanichtanankul
Synchrotron Light Research Institute (Public Organization), Nakhon Ratchasima, Thailand.
Chomphunuch
C
Songsiriritthigul
Synchrotron Light Research Institute (Public Organization), Nakhon Ratchasima, Thailand.
Kanjana
K
Thumanu
Life Science Group, Scientific Research Division, National Synchrotron Radiation Research Center, Hsinchu, Taiwan.
Chun-Jung
CJ
Chen
Department of Clinical Microscopy, Faculty of Associated Medical Sciences, Khon Kaen University, Khon Kaen, Thailand.|Cardiovascular Research Group, Khon Kaen University, Khon Kaen, Thailand.
Nantarat
N
Komanasin
D002151:Calorimetry; D028481:Euphorbia / Q000201:enzymology*; D005340:Fibrinogen / Q000737:chemistry / Q000378:metabolism*; D006031:Glycosylation; D006801:Humans; D007840:Latex / Q000737:chemistry / Q000378:metabolism*; D011485:Protein Binding; D057057:Serine Proteases / Q000737:chemistry / Q000378:metabolism*
D007840:Latex; D005340:Fibrinogen; D057057:Serine Proteases
Euphorbia cf. lactea; enzyme; fibrinogenolytic; serine protease; therapeutic proteins; toxicity