Expression and Biochemical Characterization of Acidic Phospholipase
A2I from Agkistrodon acutus
LIU Xiao-Long, WU Xiang-Fu, CAI Ming-De1 and ZHOU
Yuan-Cong*
( Shanghai Institute of Biochemistry, the Chinese Academy of Sciences,
Shanghai 200031, China；1 Shanghai Hypertension
Institute, Shanghai 200025, China )
Abstract A cDNA encoding acidic
phospholipase A2I(A.aAPLA2I)from Agkistrodon acutus
was inserted into a bacterial expression vector and effectively expressed in E.coli
RR1. The protein was produced as insoluble inclusion bodies. After partial
purification by washing the inclusion bodies with Triton X-100, denaturing and
refolding, the renatured recombinant protein was purified by FPLC column
SuperoseTM12. The enzymatic acti-vity and platelet aggregation
inhibiting effect of the expressed A.aAPLA2I is close to those of
denatured-refolded native acidic PLA2 from Agkistrodon halys
Pallas, and has the same hemolytic activity as denatured-refolded basic
phospholipase A2 from Agkistrodon halys Pallas. The roles
of various amino acid residues in the enzymatic activity and pharmacological
activities of phospholipase A2 are discussed.
Key Words Agkistrodon acutus； APLA2I； activity assay； structure； function
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