Purification and structural analysis of a murine chemotactic cytokine (CP-10) with sequence homology to S100 proteins.
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.
Lackmann
M
M
Heart Research Institute, Camperdown, New South Wales, Australia.
Cornish
C J
CJ
Simpson
R J
RJ
Moritz
R L
RL
Geczy
C L
CL
D000595:Amino Acid Sequence; D000818:Animals; D002478:Cells, Cultured; D002630:Chemotactic Factors / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification*; D002851:Chromatography, High Pressure Liquid; D002852:Chromatography, Ion Exchange; D003488:Cyanogen Bromide / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D051379:Mice; D008969:Molecular Sequence Data; D009418:S100 Proteins / Q000235:genetics*; D012689:Sequence Homology, Nucleic Acid; D013154:Spleen / Q000166:cytology; D014357:Trypsin / Q000378:metabolism
D002630:Chemotactic Factors; D009418:S100 Proteins; D014357:Trypsin; D003488:Cyanogen Bromide