Regulation and Function of 5′-Nucleotidases

Summary

Mammalian organs contain at least three different 5′-nucleotidases which favor AMP as substrate over other nucleotides by the criterion of V_max/K_m. These enzymes occur in plasma membrane, cytosol, and lysosomes. The plasma membrane enzyme has been purified to homogeneity, and its kinetic properties have been described in some detail. The enzyme shows a substantial preference for AMP over IMP [25].

Cytosolic 5′-nucleotidase from rabbit heart exhibits sigmoidal AMP saturation curves in the absence of activators. The enzyme is activated by ATP, which increases the apparent affinity of the enzyme for AMP without a change in V_max. At saturating ATP, the AMP saturation curve becomes hyperbolic in shape. The cytosolic enzyme can be dissociated and separated into catalytic and regulatory proteins. The separated catalytic protein has a hyperbolic AMP saturation curve, i. e., it exhibits Michaelis-Menten kinetics. This form is slightly inhibited by ATP. When the catalytic and regulatory proteins are mixed together, the original regulatory behavior is restored.

A lysosomal 5′-nucleotidase has been partially purified from rat liver. This enzyme too shows a preference for AMP over other nucleoside monophosphates; however, it is inhibited by ATP. The lysosomal enzyme differs in kinetic properties from various other lysosomal phosphatases and 5′-nucleotidases that have been described previously; in particular, it has no activity with 2′-and 3′-adenosine monophosphates. It shows some activity towards p-nitrophenyl phosphate, but with a much lower V_max than AMP.

Polyclonal antibody to ecto-5′-nucleotidase causes complete inhibition of the ectoenzyme, but has no effect on the cytosolic and lysosomal 5′-nucleotidases. The function of the three types of 5′-nucleotidase is discussed in terms of their localization.