Functional expression of soluble ICAM-1 by baculovirus-infected Sf9 cells.
Inflammation, metastasis and ischemia are processes that require lymphocyte or leukocyte cell recognition and adherence to endothelial counter receptors such as ICAM-1. Mapping the sites of interaction of ICAM-1 with LFA-1, the receptor for ICAM-1 on lymphocytes, may lead to the design of novel inhibitors of inflammation or metastasis. To this end, recombinant soluble ICAM-1 cDNA was engineered into the baculovirus expression system, which is capable of expressing large amounts of proteins. These constructs were designed to contain a protein leader sequence so that the transfected insect cells would secrete the recombinant polypeptide into the culture media for ease of isolation. We engineered four constructs of ICAM-1 into the baculovirus system and obtained relatively high expression of two soluble forms of ICAM-1, a two domain and a five domain form. These truncated proteins were isolated and shown to promote adherence of HL-60 cells and Molt-4 cells. These recombinant soluble proteins also inhibited cell adherence to purified intact ICAM-1 isolated from K562 cells.
Center for Life Sciences and Toxicology, Research Triangle Institute, Research Triangle Park, NC 27709.
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RR
Cobb
J S
JS
Dubins
J
J
Warner
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Molony
D000595:Amino Acid Sequence; D000818:Animals; D016367:Baculoviridae / Q000235:genetics*; D001483:Base Sequence; D002448:Cell Adhesion / Q000187:drug effects*; D015815:Cell Adhesion Molecules / Q000096:biosynthesis / Q000235:genetics* / Q000302:isolation & purification; D002460:Cell Line; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics; D005822:Genetic Vectors; D006801:Humans; D007313:Insecta; D018799:Intercellular Adhesion Molecule-1; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000302:isolation & purification / Q000494:pharmacology; D014162:Transfection
D015815:Cell Adhesion Molecules; D009838:Oligodeoxyribonucleotides; D011994:Recombinant Proteins; D018799:Intercellular Adhesion Molecule-1