Document Detail


Cloning, expression and biochemical characterization of a basic-acidic hybrid phospholipase A2-II from Agkistrodon halys pallas.
MedLine Citation:
PMID:  10209288     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
A cDNA encoding a basic-acidic hybrid phospholipase A2-II from Agkistrodon halys Pallas with an N-terminus highly homologous to that of BPLA2 and a C-terminus sequence almost the same as that of APLA2 was inserted into a bacterial expression vector and effectively expressed in Escherichia 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 superose 12. The purified recombinant enzyme with an isoelectric point of pH 6.8 could cross-react with antiserum prepared against acidic phospholipase A2. The enzymatic activity of the expressed basic-acidic hybrid phospholipase A2-II is close to that of denatured-refolded native basic phospholipase A2, and has the same inhibiting effect on platelet aggregation as denatured-refolded acidic phospholipase A2, but lacks the hemolytic activity of denatured-refolded basic phospholipase A2. To study the structural relationships among basic phospholipase A2, acidic phospholipase A2 and basic-acidic hybrid phospholipase A2-II, molecular modeling of basic-acidic hybrid phospholipase A2-II was done. The roles of various amino acid residues in the enzymatic activity and pharmacological activities of phospholipase A2 are discussed.
Authors:
X Liu; H Pan; G Yang; X Wu; Y Zhou
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Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't    
Journal Detail:
Title:  Biochimica et biophysica acta     Volume:  1431     ISSN:  0006-3002     ISO Abbreviation:  Biochim. Biophys. Acta     Publication Date:  1999 Apr 
Date Detail:
Created Date:  1999-06-01     Completed Date:  1999-06-01     Revised Date:  2007-11-15    
Medline Journal Info:
Nlm Unique ID:  0217513     Medline TA:  Biochim Biophys Acta     Country:  NETHERLANDS    
Other Details:
Languages:  eng     Pagination:  157-65     Citation Subset:  IM    
Affiliation:
Shanghai Institute of Biochemistry, Chinese Academy of Science, Shanghai 200031, China.
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MeSH Terms
Descriptor/Qualifier:
Agkistrodon*
Amino Acid Sequence
Animals
Base Sequence
Cloning, Molecular
Crotalid Venoms / enzymology*
DNA, Complementary / biosynthesis,  chemistry
Group II Phospholipases A2
Molecular Sequence Data
Phospholipases A / biosynthesis,  chemistry,  genetics*
Phospholipases A2
Chemical
Reg. No./Substance:
0/Crotalid Venoms; 0/DNA, Complementary; EC 3.1.1.-/Phospholipases A; EC 3.1.1.4/Group II Phospholipases A2; EC 3.1.1.4/Phospholipases A2

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