Molecular Design, Cloning, Expression and Purification of NH(2)-terminal Mutant of Staphylokinase deltaNSak
The 3-D structure of human microplasminogen was predicted by the method of homology modeling. The binding sites of staphylokinase and microplasminogen were determined by high-resolution genetic docking. This model is consistent with several known experimental properties of staphylokinase. To elucidate the function of NH(2)-terminus of staphylokinase for further rational reconstruction, a NH(2)-terminal-deletion mutant of staphylokinase(deltaNSak) was designed. The soluble deltaNSak was achieved with fibrinolytic activity. deltaNSak was purified by two-step ion exchange chromatography. The purity was over 95% and the specific activity of deltaNSak was 9x10(4) IU/mg. These data suggest that the 15 NH(2)-terminal amino acids of staphylokinase are not required for its plasminogen activating potential and the computer model is reasonable.
Song
Gang
G
Department of Molecular Genetics, School of Basic Medical Sciences, Shanghai Medical University, Shanghai 200032, China. hysong@shmu.edu.cn
Mo
Wei
W
Song
Hou-Yan
HY