Molecular Design,
Cloning, Expression and Purification of NH2-terminal Mutant of
Staphylokinase（ΔNSak）
SONG Gang, MO Wei and SONG Hou-Yan*
( Department of Molecular Genetics, School of Basic Medical Sciences,
Shanghai Medical University,Shanghai 200032, China )
Abstract 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 NH2-terminus
of staphylokinase for further rational reconstruction, a NH2-terminal-deletion
mutant of staphylokinase(ΔNSak) was designed . The soluble ΔNSak was achieved
with fibrinolytic activity. ΔNSak was purified by two-step ion exchange
chromatography. The purity was over 95% and the specific activity of ΔNSak was
9×104 IU/mg. These data suggest that the 15 NH2-terminal
amino acids of staphylokinase are not required for its plasminogen activating
potential and the computer model is reasonable.
Key Words staphylokinase； molecular design； mutant； expression； purification
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