THE EFFECT OF REGULATORY CA2-SITU STRUCTURES OF TROPONIN-C AND TROPONIN-I - A NEUTRON-SCATTERING STUDY( ON THE IN)

The effects of regulatory amounts of Ca2+ on the in situ structures of troponin C (TnC) and troponin I (TnI) in whole troponin have been inv estigated by neutron scattering. Ln separate difference experiments, 9 7% deuterated TnC and TnI within whole troponin were studied +/-Ca2+ i n 41.6% (H2O)-H-2 buffers in which protonated subunits were rendered ' 'invisible''. We found that the radius of gyration (R-g) Of TnI decrea sed by approximate to 10% upon addition of regulatory Ca2+ indicating that it was significantly more compact in the presence of Ca2+. The ap parent cross-sectional radius of gyration (R-c) of TnI increased by ab out 9% when regulatory Ca2+ was bound to TnC. Modeling studies showed that the high-Q scattering patterns of TnI could be fit by a TnI which consisted of two subdomains: one, a highly oblate ellipsoid of revolu tion containing about 65% of the mass and the other,a highly prolate e llipsoid of resolution consisting of about 35% of the mass. No other f its could be found with this class of models. Best fits were achieved when the axes of revolution of these ellipsoids were steeply inclined with respect to each other. Ca2+ addition decreased the center of mass separation by about 1.5 nm. The R-g of TnI, its high-Q scattering pat tern, and the resultant structure were different from previous results on neutron scattering by TnI in the (+Ca2+) TnC TnI complex. The R-g of TnC indicated that it was elongate in situ. The R-g of TnC was not sensitive to the Ca2+ occupancy of its regulatory sites. However, R-c increased upon Ca2+ addition in concert with expectations from NMR and crystallography of isolated TnC. The present observations indicate th at TnI acts like a molecular switch which is controlled by smaller Ca2 + induced changes in TnC. (C) 1998 Academic Press.