OPTICAL-ACTIVITY OF A NUCLEOTIDE-SENSITIVE TRYPTOPHAN IN MYOSIN SUBFRAGMENT-1 DURING ATP HYDROLYSIS

The xanthene probes 5'-iodoacetamido-fluorescein and -tetramethylrhoda mine specifically modify skeletal muscle myosin subfragment 1 (S1) at the reactive thiol residue (SH1) and fully quench the fluorescence emi ssion from tryptophan residue 510 (Trp510) in S1 (T.P. Burghardt and K . Ajtai, Biophys. Chem., 60 (1996) 119; K. Ajtai and T.P. Burghardt, B iochemistry, 34 (1995) 15943), The difference between the fluorescence intensity obtained from S1 and probe-modified S1 comes solely from Tr p510 in chymotryptic S1, a protein fragment that contains five tryptop han residues. The rotary strength and quantum efficiency of Trp510 wer e measured using difference signals from fluorescence detected circula r dichroism (FDCD) and fluorescence emission spectroscopy. These struc ture-sensitive signals indicate that the binding of nucleotide or nucl eotide analogs to the active site of S1 causes structural changes in S 1 at Trp510 and that a one-to-one correspondence exists between Trp510 conformation and transient states of myosin during contraction, The T rp510 rotary strength and quantum efficiency were interpreted structur ally in terms of the indole side-chain conformation using model struct ures and established computational methods.