Circular dichroic spectra of 6-thioguanosine nucleotides and their complexes with myosin subfragment 1

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Abstract
The circular dichroic [CD] spectra of the thione form of 2-amino-6-mercapto-9.beta.-ribofuranosylpurine 5''-triphosphate (thioGTP), thioGDP, thioGTP(.gamma.S), thioGMP-P(NH)P and 6-mercapto-9.beta.-ribofuranosylpurine 5''-diphosphate (thioIDP) and their complexes with [rabbit muscle] myosin subfragment 1 and heavy meromyosin were measured between 300 and 400 nm. The free nucleotides have a weak negative CD spectral peak at their absorption maxima, with a longer wavelength shoulder. On binding to the proteolytic fragments of myosin, the negative peak is enhanced .apprx. 10-fold and a longer wavelength positive peak appears. These effects are attributed to a change in the stereochemical structure of the nucleotide and specific interactions within the nucleotide binding site. All 4 thioguanosine nucleotides give these spectral changes, although minor significant differences occur. The CD spectra of the subfragment 1 steady-state complexes with thioGTP and thioGTP(.gamma.S) are similar to each other but different from the complexes of subfragment 1 with thioGDP and thioGMP-P(NH)P. Interactions between nucleotides in the complexes are excluded by a study of their spectra in 2-propanol when base stacking occurs. This is the 1st investigation where CD was used to determine structural differences between different myosin-nucleotide states and provides evidence that the nucleotide in the .gamma.S bound state is similar to that of the triphosphate bound state.