Ultraviolet difference spectroscopic studies of AMP and dAMP upon binding to snake muscle fructose 1, 6-bisphosphatase showed that both of the ligands interacting with the enzyme generated three positive peaks located at 270 nm, 280 nm and 288 nm. The binding constants calculated from the titration curve of difference spactra by the successive addition of nueleotides were 2.0×10~6M~(-1) and 1.2×10~6M~(-1) respectively. The fact of the same binding constants for the two ligands indicated that the unique inhibition behavior of dAMP differing from AMP was not due to the difference of binding between AMP and dAMP, but due to the absence of 2’-OH group on the ribose of dAMP. In contrast to the difference spectra of AMP binding, the U. V. difference spectra produced by tryptic digestion of fructose 1, 6-bisphosphatase gave two negative bands at 280 nm and 288 nm and had a close resemblance to that of the enzyme in urea. The opposite signs of the two kinds of difference U. V. spectra produced by AMP binding Ultraviolet difference spectroscopic studies of AMP and dAMP upon binding to snake muscle fructose 1, 6-bisphosphatase showed that both of the ligands interact with the enzyme generated three positive peaks located at 270 nm, 280 nm and 288 nm. The binding constants calculated from the titration curve of difference spactra by the successive addition of nueleotides were 2.0 × 10 -6 M -1 and 1.2 × 10 -6 M -1 respectively. The fact of the same binding constants for the two ligands that that the unique inhibition behavior of dAMP differing from AMP was not due to the difference of binding between AMP and dAMP, but due to absence of 2’-OH group on the ribose of dAMP. In contrast to the difference spectra of AMP binding, the UV difference spectra produced by tryptic digestion of fructose 1, 6-bisphosphatase gave two negative bands at 280 nm and 288 nm and had a close resemblance to that of the enzyme in urea. The opposite signs of the two kinds of difference UV spectra produc ed by AMP binding