کد مقاله کد نشریه سال انتشار مقاله انگلیسی نسخه تمام متن
5371388 1388818 2011 8 صفحه PDF دانلود رایگان
عنوان انگلیسی مقاله ISI
Rigidification of the autolysis loop enhances Na+ binding to thrombin
موضوعات مرتبط
مهندسی و علوم پایه شیمی شیمی تئوریک و عملی
پیش نمایش صفحه اول مقاله
Rigidification of the autolysis loop enhances Na+ binding to thrombin
چکیده انگلیسی

Binding of Na+ to thrombin ensures high activity toward physiological substrates and optimizes the procoagulant and prothrombotic roles of the enzyme in vivo. Under physiological conditions of pH and temperature, the binding affinity of Na+ is weak due to large heat capacity and enthalpy changes associated with binding, and the Kd = 80 mM ensures only 64% saturation of the site at the concentration of Na+ in the blood (140 mM). Residues controlling Na+ binding and activation have been identified. Yet, attempts to improve the interaction of Na+ with thrombin and possibly increase catalytic activity under physiological conditions have so far been unsuccessful. Here we report how replacement of the flexible autolysis loop of human thrombin with the homologous rigid domain of the murine enzyme results in a drastic (up to 10-fold) increase in Na+ affinity and a significant improvement in the catalytic activity of the enzyme. Rigidification of the autolysis loop abolishes the heat capacity change associated with Na+ binding observed in the wild-type and also increases the stability of thrombin. These findings have general relevance to protein engineering studies of clotting proteases and trypsin-like enzymes.

Research highlights► Replacement of the flexible autolysis loop of human thrombin with the homologous rigid domain of the murine enzyme results in a drastic (up to 10-fold) increase in Na+ affinity and a significant improvement in the catalytic activity of the enzyme. ► Rigidification of the autolysis loop abolishes the heat capacity change associated with Na+ binding observed in the wild-type and also increases the stability of the enzyme.

ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Biophysical Chemistry - Volume 159, Issue 1, November 2011, Pages 6-13
نویسندگان
, , , , ,