Identification of a novel eosinophil chemotactic cytokine (ECF-L) as a chitinase family protein

A novel eosinophil chemotactic cytokine (ECF-L) was purified from the cultu re supernatant of splenocytes of mice by a combination of anion-exchange ch romatography, Procion red-agarose affinity chromatography, size exclusion h igh performance liquid chromatography (HPLC), and reverse phase HPLC. The N H2-terminal amino acid sequence was determined by direct protein sequencing . An ECF-L cDNA clone of 1,506 nucleotides was isolated from a cDNA library , and the nucleotide sequence predicted a mature protein of 397 amino acids . A recombinant ECF-L showed a level of eosinophil chemotactic activity com parable with that of natural ECF-L, and the activity was inhibited by a mon oclonal antibody to ECF-L. ECF-L also attracted T lymphocytes and bone marr ow polymorphonuclear leukocytes in vitro, whereas it caused selective extra vasation of eosinophils in vivo. ECF-L mRNA was highly expressed in spleen, bone marrow, lung, and heart. A comprehensive GenBank data base search rev ealed that ECF-L is a chitinase family protein, ECF-L retains those amino a cids highly conserved among chitinase family proteins, but Asp and Glu resi dues essential for the proton donation in hydrolysis were replaced by Asn a nd Gin, respectively. Although ECF-L contains a consensus CXC sequence near the NH, terminus akin to chemokine family proteins, the rest of ECF-L show s poor homology with chemokines.