Purification of granulocyte chemotactic peptide/interleukin-8 reveals N-terminal sequence heterogeneity similar to that of beta-thromboglobulin.
Stimulated human peripheral blood leukocytes produce a chemotactic factor for granulocytes (granulocyte chemotactic peptide/interleukin-8; GCP/IL-8), which is structurally related to platelet-derived beta-thromboglobulin. Analytically pure CGP/IL-8 and beta-thromboglobulin could be obtained after three purification steps, comprising adsorption to silicic acid, heparin-Sepharose chromatography and ion-exchange chromatography. Although GCP/IL-8 and beta-thromboglobulin had a similar affinity for heparin, they could be separated on a cation-exchange column. Both molecules were heterogeneous in that 6-7-kDa protein doublets were detected upon SDS/PAGE. N-terminal amino acid sequence analysis revealed the presence of six immunologically related but differently truncated polypeptides of beta-thromboglobulin, of which only two corresponded to previously described forms. Similarly, apart from a major polypeptide, five minor species of GCP/IL-8 were detected that also differed by N-terminal truncation. The most processed forms of beta-thromboglobulin and GCP/IL-8 were found to have their N-terminus in that region of the primary structure where a significant similarity between the two molecules starts. GCP/IL-8 was found to be chemotactic for granulocytes with a specific activity of 10(5) units/mg, whereas none of the beta-thromboglobulin species possessed detectable chemotactic activity.
Van Damme
J
J
Rega Institute for Medical Research, University of Leuven, Belgium.
Van Beeumen
J
J
Conings
R
R
Decock
B
B
Billiau
A
A
D000595:Amino Acid Sequence; D002630:Chemotactic Factors / Q000097:blood / Q000235:genetics* / Q000302:isolation & purification; D002634:Chemotaxis, Leukocyte; D002846:Chromatography, Affinity; D002852:Chromatography, Ion Exchange; D004591:Electrophoresis, Polyacrylamide Gel; D006098:Granulocytes / Q000502:physiology*; D006801:Humans; D016209:Interleukin-8; D007378:Interleukins / Q000097:blood / Q000235:genetics* / Q000302:isolation & purification; D007962:Leukocytes / Q000502:physiology*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D012689:Sequence Homology, Nucleic Acid; D001620:beta-Thromboglobulin / Q000235:genetics* / Q000302:isolation & purification
D002630:Chemotactic Factors; D016209:Interleukin-8; D007378:Interleukins; D001620:beta-Thromboglobulin