Further Studies on the Buffer-Catalyzed Cleavage and Isomerization of Uridyluridine. Medium and Ionic Strength Effects on Catalysis by Morpholine, Imidazole, and Acetate Buffers Help Clarify the Mechanisms Involved and Their Relationship to the Mechanism Used by the Enzyme Ribonuclease and by a Ribonuclease Mimic

The cleavage and isomerization of 3‘,5‘-uridyluridine catalyzed by
morpholine buffers and by imidazole
buffers has been reinvestigated. The key evidence for a previously
proposed partitioning mechanismin which the
buffer acid BH+ converts the substrate to a phosphorane
monoanion intermediate which then partitions either to the
2‘,5‘ isomer of the substrate or (with buffer base catalysis) to the
cleavage productis confirmed. The negative
catalytic effect of the buffer base on the isomerization reaction is
not due to a medium effect. Indeed the medium
effect on this reaction is in the opposite direction, strengthening the
catalytic evidence. However, this branching
mechanism with sequential bifunctional catalysis of the cleavage
reaction is accompanied by an additional cleavage
path using the buffer base only. This additional path, for which
several alternative mechanisms are possible, is
required by the results of improved studies on the imidazole catalysis.
These show that the previously reported
decrease in rate at a low BH+/B ratio is due to ionic
strength effects. The relative importance of these
two
pathwaysone with kinetic dependence on the buffer acid and one
without such dependencedepends on the buffer
basicity/acidity. With the acidic buffer acetate/acetic acid, a
buffer-acid-catalyzed mechanism for the cleavage and
the isomerization is dominant. A bifunctional mechanism, in which
one step involves simultaneous acid−base catalysis,
seems most likely. The medium effects of added dioxane on all
these reactions are sensible in terms of the detailed
mechanisms proposed. The relationship of these results to the
mechanisms of catalysis by ribonuclease and by an
enzyme mimic is discussed.