Incorporation of low molecular weight molecules into alpha(2)-macroglobulin by nucleophilic exchange.
alpha(2)-Macroglobulin (alpha(2)M) is a proteinase inhibitor that functions by a trapping mechanism which has been exploited such that the receptor-recognized, activated form (alpha(2)M( *)) can be employed to target antigens to antigen-presenting cells. Another potential use of alpha(2)M( *) is as a drug delivery system. In this study we demonstrate that guanosine triphosphate, labeled with Texas red (GTP-TR) formed complexes with alpha(2)M( *) following activation by proteolytic or non-proteolytic reactions. Optimal incorporation occurred with 20 microM GTP-TR, pH 8.0 for 5h at 50 degrees C. NaCl concentration (100 or 200 mM) had little effect on incorporation at this pH or temperature, but was significant at sub-optimum temperature and pH values. Maximum incorporation was 1.2 mol GTP-TR/mol alpha(2)M( *). PAGE showed that 70-90% of the GTP-TR is bound in a SDS/2-mercaptoethanol resistant manner. Guanosine, adenosine, and imidazole competed with GTP-TR to form complexes with alpha(2)M( *).
Bond
Jennifer E
JE
Department of Pathology, Duke University Medical Center, Box 3712, Durham, NC 27710, USA.
Cianciolo
George J
GJ
Pizzo
Salvatore V
SV
D001665:Binding Sites; D008970:Molecular Weight; D009705:Nucleosides / Q000737:chemistry*; D011485:Protein Binding; D015202:Protein Engineering / Q000379:methods; D013194:Staining and Labeling / Q000379:methods*; D000511:alpha-Macroglobulins / Q000737:chemistry*
D009705:Nucleosides; D000511:alpha-Macroglobulins