Calcium-dependent complex assembly of the myeloic differentiation proteins MRP-8 and MRP-14.
MRP-8 and MRP-14 are calcium-binding proteins belonging to the S-100 protein family which have been shown to be associated with specific stages of myeloic/monocytic cell differentiation. Members of this protein family are shown to form homo- and heterodimers. Complex formation has also been observed in preliminary experiments for MRP-8 and MRP-14. To evaluate the in vivo relevance of the MRP complex formation and the stoichiometric ratio of individual components complexes were isolated from granulocytes and monocytes by immunoaffinity chromatography using monospecific antibodies. The purified fraction of the MRPs was found to contain monomers and dimers as shown on sodium dodecyl sulfate-polyacrylamide gel electrophoresis by silver staining and immunoblotting. Similar results were obtained by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunoblotting of crude cell extracts. The existence of the MRP complexes in vivo was demonstrated by chemical cross-linking and subsequent isolation of complexes by immunoaffinity chromatography. Two new, highly abundant complexes were found in addition to the heterodimer, but neither monomers nor homodimers were detected. The two larger protein complexes (35.0 and 48.5 kDa) were identified as [MRP-8)2.(MRP-14] trimer and [MRP-8)2.(MRP-14)2) tetramer, respectively. All complexes could be shown to be noncovalently associated in vivo. Furthermore, the association of MRPs was shown to be Ca2+ dependent.
Teigelkamp
S
S
Institute of Experimental Dermatology, University of Munster, Germany.
Bhardwaj
R S
RS
Roth
J
J
Meinardus-Hager
G
G
Karas
M
M
Sorg
C
C
D015153:Blotting, Western; D002118:Calcium / Q000494:pharmacology*; D002135:Calcium-Binding Proteins / Q000097:blood* / Q000302:isolation & purification; D040501:Calgranulin A; D040502:Calgranulin B; D002454:Cell Differentiation; D002846:Chromatography, Affinity; D004591:Electrophoresis, Polyacrylamide Gel; D006098:Granulocytes / Q000378:metabolism*; D006801:Humans; D007460:Iodoacetamide / Q000494:pharmacology; D046911:Macromolecular Substances; D008970:Molecular Weight; D009000:Monocytes / Q000378:metabolism*; D013438:Sulfhydryl Compounds / Q000032:analysis
D002135:Calcium-Binding Proteins; D040501:Calgranulin A; D040502:Calgranulin B; D046911:Macromolecular Substances; D013438:Sulfhydryl Compounds; D002118:Calcium; D007460:Iodoacetamide