Purification and structural analysis of a murine chemotactic cytokine (CP-10) with sequence homology to S100 proteins

• Published in: The Journal of biological chemistry 1992-04, Vol.267 (11), p.7499-7504
• Main Authors: LACKMANN, M, CORNISH, C. J, SIMPSON, R. J, MORITZ, R. L, GECZY, C. L
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Analysis of the immune response. Humoral and cellular immunity; Animals; Biological and medical sciences; Cells, Cultured; Chemotactic Factors - chemistry; Chemotactic Factors - genetics; Chemotactic Factors - isolation & purification; Chromatography, High Pressure Liquid; Chromatography, Ion Exchange; Cyanogen Bromide - metabolism; Electrophoresis, Polyacrylamide Gel; Fundamental and applied biological sciences. Psychology; Fundamental immunology; Immunobiology; Lymphokines, interleukins ( function, expression); Mice; Molecular Sequence Data; Regulatory factors and their cellular receptors; S100 protein; S100 Proteins - genetics; Sequence Homology, Nucleic Acid; Spleen - cytology; Trypsin - metabolism
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1016/S0021-9258(18)42545-8

In delayed-type hypersensitivity reactions, cytokine-mediated cell migration leads to localized accumulation of neutrophils and mononuclear cells over 4-48 h. In contrast to transient (2-6 h) responses elicited by other chemotactic factors, earlier studies indicated that a chemotactic activity previously described in our laboratory elicited skin test responses over 24 h, identical to those induced by injection of antigen into a sensitized test subject. We have isolated this factor, a 10.3-kDa chemotactic protein designated CP-10, from supernatants of activated murine spleen cells. Purification to homogeneity was achieved using affinity chromatography on iminodiacetic acid-immobilized copper and cation-exchange, mixed mode (cation exchange/metal affinity), reversed-phase, and size-exclusion high performance liquid chromatography. CP-10 had maximal chemotactic activity for neutrophils at 10(-13) M. The 76-amino acid sequence, obtained by automated N-terminal microsequence analysis of native CP-10, and fragments derived from trypsin digestion and cyanogen bromide cleavage indicated no sequence identity with any known cytokine or chemotactic factor but revealed up to 55% sequence homology with S100, Ca2(+)-binding proteins. CP-10 appears to be the first protein of this family with a well defined function affecting cell migration, and its biological potency suggests an important role for this cytokine in cellular immune reactions.