Browsing by Author "Lin, ES"

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Colorimetric determination of the purity of 3 '-phospho adenosine 5'-phosphosulfate and natural abundance of 3 '-phospho adenosine 5 '-phosphate at picomole quantities
(1998-11-01)
Lin, ES
;
Yang, YS
;
生物科技學系
;
Department of Biological Science and Technology
This work presents novel colorimetric methods not only to measure 3'-phospho adenosine 5'-phosphate (PAP) and 3'-phospho adenosine 5'-phosphosulfate (PAPS) in the range of picomoles, but also to determine the purity of PAPS or PAP contaminants in PAPS in the range of nanomoles. These methods exploit the availability of overexpressed phenol sulfotransferase (PST) and the fact that sulfuryl group transfer requires the use of PAP or PAPS as a cofactor or cosubstrate, Experimental results indicate that absorption at 400 nm due to the production of 4-nitrophenol (pNP) is catalyzed by PST when the sulfuryl group transfers from 4-nitrophenylsulfate (pNPS) to PAP or to 2-napthol, In the absence of an acceptor substrate, PAPS is hydrolyzed to PAP by PST and is determined by sulfation with pNPS before and after this reaction. The change of absorption of pNP at 400 nm corresponds to the amount of PAP that is hydrolyzed from PAPS. Moreover, a standard curve is constructed using authentic PAP and PAP-free PST. Furthermore, this curve is used to determine the amount of PAP in extracts of pig liver, rat liver, and Escherichia coli. (C) 1998 Academic Press.
Effects of 3 '-phosphoadenosine 5 '-phosphate on the activity and folding of phenol sulfotransferase
(1998-02-20)
Yang, YS
;
Tsai, SW
;
Lin, ES
;
生物科技學系
;
Department of Biological Science and Technology
Known spectroscopic and kinetic data are used to formulate pathways of the physiological and transfer reactions and the substrate inhibition of phenol sulfotransferase. Kinetic mechanisms indicate that release of PAP from enzyme complex is required for the physiological reaction but not for the transfer reaction. The pathways explain rate difference between the physiological and transfer reactions since the release of PAP is the rate-limiting step of the former reaction. Two enzyme species of phenol sulfotransferase which distinguish the physiological and transfer reaction were found to involve the binding of PAP. Differences between two forms of phenol sulfotransferase, alpha and beta, indicate that they assemble through different folding process. It is demonstrated that only alpha enzyme renatures in the presence of PAP and beta enzyme renatures only ir. the absence of PAP in vitro. In the over-expressed system, formation of alpha and beta phenol sulfotransferase is also dependent on the availability of PAP in Escherichia coli. It is concluded that folding of phenol sulfotransferase is assisted by PAP to form alpha enzyme. In the absence of PAT, beta form of phenol sulfotransferase is produced. (C) 1998 Elsevier Science Ireland Ltd. All rights reserved.
Modeling the binding and inhibition mechanism of nucleotides and sulfotransferase using molecular docking
(2003-06-01)
Lin, ES
;
Yang, JM
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Yang, YS
;
生物科技學系
;
生物資訊及系統生物研究所
Phenol sulfotransferase (PST) catalyzes sulfuryl group transfer from adenosine 3'-phosphate 5'-phosphosulfate (PAPS) to a variety of nucleophilic acceptors in biological systems. Physiological sulfation by PAPS results in the production of adenosine 3',5'-diphosphate (PAP). PAP may become a strong inhibitor or cofactor for the physiological or the transfer reactions, respectively. Several nucleotides other than, PAP also serve as substrates, inhibitors or cofactors of PST catalyzed reactions. We are interested in the effects of these nucleotides on the PST catalyzed reactions and their possible physiological significance in biology. Several nucleotide inhibitors (such as adenosine 5'-diphosphate and adenosine 5'-triphosphate) of sulfotransferase in physiological reactions have been reported in the literature. However, our data suggests that they are not inhibitors for the transfer reaction or the reverse physiological reaction catalyzed by PST. The aim of this work is to show that the molecular docking analysis can be successfully used to underline the inhibition mechanism of these nucleotides. First, the selected compounds were subjected to a detailed docking analysis, by means of GEMDOCK, a program able to reveal the most likely binding mode for each ligand. By comparing these results with binding sites and binding compounds of the nucleotides with known X-ray structures, it is possible to highlight the site specificity and the inhibition mechanism of the select compounds. The results obtained by the above algorithm were further confirmed experimentally. In this paper, we show the effect of a variety of nucleotides on the activity of sulfotransferase in different docking conditions.
Nucleotide binding and sulfation catalyzed by phenol sulfotransferase
(2000-05-19)
Lin, ES
;
Yang, YS
;
生物科技學系
;
Department of Biological Science and Technology
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 P-31 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. (C) 2000 Academic Press.
Nucleotides as cofactor, inhibitor, and antioxidation and stabilization reagent of phenol sulfotransferase
(2001-03-07)
Lin, ES
;
Li, TS
;
Yang, YS
;
交大名義發表
;
National Chiao Tung University
Reactions of phenol sulfotransferase with nucleotides
(1997-07-31)
Yang, YS
;
Lin, ES
;
生物科技學系
;
Department of Biological Science and Technology
Reactions of phenol sulfotransferase with nucleotides
(1997-07-31)
Yang, YS
;
Lin, ES
;
生物科技學系
;
Department of Biological Science and Technology