Cytotoxic Cell Granule-mediated Apoptosis CHARACTERIZATION OF THE MACROMOLECULAR COMPLEX OF GRANZYME B WITH SERGLYCIN

摘要: We have recently shown that the physiological mediator of granule-mediated apoptosis is a macromolecular complex granzymes and perforin complexed with chondroitin-sulfate proteoglycan, serglycin (Metkar, S. S., Wang, B., Aguilar-Santelises, M., Raja, Uhlin-Hansen, L., Podack, E., Trapani, J. A., Froelich, C. (2002) Immunity 16, 417-428). now report our biophysical studies establishing nature granzyme B-serglycin (GrB.SG) complex. Dynamic laser light scattering establish SG has hydrodynamic radius approximately 140 +/- 23 nm, comparable to some viral particles. Agarose mobility shift gels surface plasmon resonance (SPR), show binds tightly GrB capacity hold 30-60 molecules. SPR also indicate equivalent binding affinities (K(d) 0.8 microm), under acidic (granule pH) neutral isotonic conditions (extra-cytoplasmic pH), for GrB.SG interaction. Finally, characterization interactions within granules revealed complexes two distinct molecular sizes, one held 4-8 molecules GrB, whereas other contained as many 32 or granule proteins. These provide firm basis earlier reported observations proapoptotic exocytosed predominantly SG.