An insight into the thermodynamic characteristics of human thrombopoietin complexation with TN1 antibody

作者：Arai Shigeki^*; Shibazaki Chie; Adachi Motoyasu; Honjo Eijiro; Tamada Taro; Maeda Yoshitake; Tahara Tomoyuki; Kato Takashi; Miyazaki Hiroshi; Blaber Michael; Kuroki Ryota

来源：Protein Science, 2016, 25(10): 1786-1796.

DOI：10.1002/pro.2985

• TN1
• thrombopoietin
• antigen-antibody interaction
• isothermal titration calorimetry
• X-ray crystallography

Human thrombopoietin (hTPO) primarily stimulates megakaryocytopoiesis and platelet production and is neutralized by the mouse TN1 antibody. The thermodynamic characteristics of TN1 antibody-hTPO complexation were analyzed by isothermal titration calorimetry (ITC) using an antigen-binding fragment (Fab) derived from the TN1 antibody (TN1-Fab). To clarify the mechanism by which hTPO is recognized by TN1-Fab the conformation of free TN1-Fab was determined to a resolution of 2.0 angstrom using X-ray crystallography and compared with the hTPO-bound form of TN1-Fab determined by a previous study. This structural comparison revealed that the conformation of TN1-Fab does not substantially change after hTPO binding and a set of 15 water molecules is released from the antigen-binding site (paratope) of TN1-Fab upon hTPO complexation. Interestingly, the heat capacity change (Cp) measured by ITC (-1.52 +/- 0.05 kJmol(-1)K(-1)) differed significantly from calculations based upon the X-ray structure data of the hTPO-bound and unbound forms of TN1-Fab (-1.02 approximate to 0.25 kJmol(-1)K(-1)) suggesting that hTPO undergoes an induced-fit conformational change combined with significant desolvation upon TN1-Fab binding. The results shed light on the structural biology associated with neutralizing antibody recognition.

• 出版日期2016-10