PMID: 9548595Apr 21, 1998Paper

Identification of the epitope of a monoclonal antibody that inhibits heparin binding of lipoprotein lipase: new evidence for a carboxyl-terminal heparin-binding domain.

Journal of Lipid Research
R A SendakAndré Bensadoun

Abstract

A panel of 13 monoclonal antibodies to avian lipoprotein lipase (LPL) was screened for inhibition of LPL binding to primary avian adipocytes. One monoclonal antibody, designated xCAL (monoclonal antibody to chicken adipose lipoprotein lipase) 3-6a, was found to inhibit the binding of LPL to primary avian adipocytes. In solid phase assays, xCAL 3-6a inhibited the binding of LPL to both heparan sulfate and heparin. XCAL 3-6a did not inhibit the catalytic activity of the avian enzyme. The monoclonal antibody was not found to cross-react significantly with bovine lipoprotein lipase. In order to determine the location of the epitope of xCAL 3-6a on lipoprotein lipase, several avian lipoprotein lipase deletion mutants were constructed and produced as glutathione S-transferase (GST) fusion proteins in E. coli. These mutants were screened for their ability to react with xCAL 3-6a using Western blotting. The minimum continuous fragment of lipoprotein lipase that was required for reactivity contained the amino acids 310 to 450. Site-directed mutagenesis of basic residues 321, 405, 407, 409, 415, and 416 revealed that Arg 405 is necessary for the interaction of LPL with xCAL 3-6a. Additional deletions of either the amino- or carboxyl-term...Continue Reading

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