Lin E S, Yang Y S
Department of Biological Science and Technology, College of Science, Hsinchu, Taiwan, Republic of China.
Biochem Biophys Res Commun. 2000 May 19;271(3):818-22. doi: 10.1006/bbrc.2000.2722.
The sulfation of a nucleotide is an indispensable step for the sulfuryl group transfer in a biological system. The product and cosubstrate of sulfotransferase in physiological condition are adenosine 3',5'-bisphosphate (PAP) and 3'-phospho adenosine 5'-phosphosulfate (PAPS), respectively. We find that ribose and adenine, two major parts of the adenosine nucleotide, bind tightly to phenol sulfotransferase (PST) separately, and various nucleotides also bind tightly to PST. We determine the dissociation constants of a variety of nucleotides and examine their potential as cofactors or cosubstrates of PST. Using 4-nitrophenyl sulfate as the sulfuryl group donor, three nucleotides, 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 position by a PST catalyzed reaction. Spectrophotometry, HPLC, and (31)P NMR are used 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 existence and possible physiological significance of these newly reported binding and sulfation of nucleotides by PST in biology is yet to be discovered.
核苷酸的硫酸化是生物系统中硫酰基转移不可或缺的一步。在生理条件下，磺基转移酶的产物和共底物分别是3',5'-二磷酸腺苷（PAP）和3'-磷酸腺苷5'-磷酸硫酸酯（PAPS）。我们发现，腺苷核苷酸的两个主要部分——核糖和腺嘌呤，分别与酚磺基转移酶（PST）紧密结合，并且各种核苷酸也与PST紧密结合。我们测定了多种核苷酸的解离常数，并研究了它们作为PST辅因子或共底物的潜力。以硫酸对硝基苯酯作为硫酰基供体，首次证明了三种核苷酸，即5'-单磷酸腺苷（AMP）、2',5'-二磷酸腺苷（2',5'-PAP）和2':3'-环磷酸腺苷5'-磷酸酯（2':3'-环PAP），在PST催化的反应中于5'-磷酸位置发生硫酸化。采用分光光度法、高效液相色谱法和³¹P核磁共振法测定PST的活性并鉴定硫酸化的核苷酸。当这些核苷酸用作硫酰基转移的辅因子或共底物时，测定了PST的V(max)和这些核苷酸的K(m)。PST对核苷酸的这些新报道的结合和硫酸化在生物学中的存在及其可能的生理意义还有待发现。
