Optical activity of a nucleotide-sensitive tryptophan in myosin subfragment 1 during ATP hydrolysis.
The xanthene probes 5'-iodoacetamido-fluorescein and -tetramethylrhodamine specifically modify skeletal muscle myosin subfragment 1 (S1) at the reactive thiol residue (SH1) and fully quench the fluorescence emission from tryptophan residue 510 (Trp510) in S1 (T.P. Burghardt and K. Ajtai, Biophys. Chem., 60 (1996) 119; K. Ajtai and T.P. Burghardt, Biochemistry, 34 (1995) 15943). The difference between the fluorescence intensity obtained from S1 and probe-modified S1 comes solely from Trp510 in chymotryptic S1, a protein fragment that contains five tryptophan residues. The rotary strength and quantum efficiency of Trp510 were measured using difference signals from fluorescence detected circular dichroism (FDCD) and fluorescence emission spectroscopy. These structure-sensitive signals indicate that the binding of nucleotide or nucleotide analogs to the active site of S1 causes structural changes in S1 at Trp510 and that a one-to-one correspondence exists between Trp510 conformation and transient states of myosin during contraction. The Trp510 rotary strength and quantum efficiency were interpreted structurally in terms of the indole side-chain conformation using model structures and established computational methods.
Park
S
S
Department of Biochemistry and Molecular Biology, Mayo Foundation, Rochester, MN 55905, USA.
Ajtai
K
K
Burghardt
T P
TP
D000255:Adenosine Triphosphate / Q000378:metabolism*; D000818:Animals; D001665:Binding Sites; D002645:Chickens; D002942:Circular Dichroism; D018360:Crystallography, X-Ray; D005452:Fluoresceins / Q000378:metabolism; D005456:Fluorescent Dyes / Q000378:metabolism; D008958:Models, Molecular; D018482:Muscle, Skeletal / Q000737:chemistry; D015879:Myosin Subfragments / Q000378:metabolism*; D009711:Nucleotides / Q000378:metabolism; D009904:Optical Rotation; D011817:Rabbits; D012235:Rhodamines / Q000378:metabolism; D013050:Spectrometry, Fluorescence; D013053:Spectrophotometry; D013438:Sulfhydryl Compounds / Q000378:metabolism; D014364:Tryptophan / Q000378:metabolism*
D005452:Fluoresceins; D005456:Fluorescent Dyes; D015879:Myosin Subfragments; D009711:Nucleotides; D012235:Rhodamines; D013438:Sulfhydryl Compounds; D014364:Tryptophan; D000255:Adenosine Triphosphate