Chemical modification of tropomyosin with NBD-chloride

Muscle tropomyosin was modified with 7-chloro-4-nitrobenzo-2-oxa-1,3-diazole (NBD-chloride) at several different pH values. NBD-chloride reacts specifically with SH residue at neutral pH but it reacts with both SH residue and amino residues at alkaline pH. The polymerizability of tropomyosin at low ionic strength and the binding property of tropomyosin to F-actin were not affected by the modification of SH residues but they were lost rapidly by the modification of amino groups, in accordance with the previous report [Johnson, P. & Smillie, L.B. (1977) Biochemistry 16, 2264-2269]. By the addition of heavy meromyosin, labeled tropomyosin which could not bind to F-actin recovered the binding ability to F-actin and it could regulate the superprecipitation of actomyosin in the presence of troponin. Further modification of amino groups (labeling ratios more than 5) led to loss of the regulating ability completely.