The sulfation of a nucleotide is an indispensable step for the sulfuryl gro
up transfer in a biological system. The product and cosubstrate of sulfotra
nsferase in physiological condition are adenosine 3',5'-bisphosphate (PAP)
and 3'-phospho adenosine 5'-phosphosulfate (PAPS), respectively. We find th
at ribose and adenine, two major parts of the adenosine nucleotide, bind ti
ghtly to phenol sulfotransferase (PST) separately, and various nucleotides
also bind tightly to PST. We determine the dissociation constants of a vari
ety of nucleotides and examine their potential as cofactors or cosubstrates
of PST. Using 4-nitrophenyl sulfate as the sulfuryl group donor, three nuc
leotides, adenosine 5'-monophosphate (AMP), adenosine 2',5'-bisphosphate (2
',5'-PAP), and adenosine 2':3' cyclic phosphate 5'-phosphate (2':3'-cyclic
PAP), are shown here for the first time to be sulfated at 5'-phopho positio
n by a PST catalyzed reaction. Spectrophotometry, HPLC, and P-31 NMR are us
ed to determine the activity of PST and identify the sulfated nucleotides.
The V-max of PST and K-m of these nucleotides are determined when they are
used as cofactors or cosubstrates for the sulfuryl group transfer. The exis
tence and possible physiological significance of these newly reported bindi
ng and sulfation of nucleotides by PST in biology is yet to be discovered.
(C) 2000 Academic Press.