JBC, Vol. 252, Issue 6, 2089-2094, Mar, 1977
Inhibition and inactivation of bovine mammary and liver UDP-galactose-4-epimerases
C. R. Geren, L. M. Geren and K. E. Ebner
Bovine liver and mammary UDP-galactose-4-epimerases were investigated with
respect to various inhibitors and inactivators. Uridine nucleotides and
NADH are potent inhibitors with Ki values in the low micromolar range. The
NAD+/NADH ratio may be an important physiological control mechanism for it
affects markedly the activity of the enzyme with 50% inhibition occurring
at a ratio of 20:1. In the presence of uridine nucleotides binding of NADH
to the epimerases is enhanced. Consequently, the effect of changes in the
NAD+/NADH ratio in vivo would not be immediately apparent as uridine
nucleotides would slow down the displacement of NADH by NAD+. Neither
uridine nor galactose 1-phosphate inhibits the purified enzymes as
previously reported with the impure liver enzyme. Uridine nucleotides
provide almost total protection against the apparent first order
inactivation of the epimerases by trypsin and allow determination of
dissociation constants. NAD+ partially protects against trypsin
inactivation. Inactivation with various sulfhydryl reagents is complex and
the results indicate that at least three sulfhydryl groups may be modified
before total inactivation occurs. Partial inactivation occurs upon
modification of the epimerases with 2-hydroxy-5-nitrogenzyl bromide. Some
protection against this modification is provided by the combination of NAD+
and UDP.
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