当前位置： SCI文献检索 > ARCHIVES OF BIOCHEMISTRY AND BIOPHYSICS期刊下所有文献 > Synthesis and properties of adenosine-5'-triphosphoro-gamma-1-(5-sulfonic acid)naphthyl ethylamidate: a fluorescent nucleotide substrate for DNA-dependent RNA polymerase from Escherichia coli.

Synthesis and properties of adenosine-5'-triphosphoro-gamma-1-(5-sulfonic acid)naphthyl ethylamidate: a fluorescent nucleotide substrate for DNA-dependent RNA polymerase from Escherichia coli.

Abstract：

:A new fluorescent ATP analog, adenosine-5'-triphosphoro-gamma-1-(5-sulfonic acid)naphthyl ethylamidate (gamma-1,5-EDANS)ATP, containing the fluorophore N-(aminoethyl)-5-naphthylamine-1-sulfonic acid attached via a gamma-phosphoamidate bond was synthesized in good yield. It has absorption maxima at 255 and 344 nm and a fluorescence emission maximum at 490 nm. These spectral characteristics permit its uses as an energy acceptor for energy transfer from the intrinsic protein fluorophores and as an energy donor for the energy transfer to the intrinsic Co of Co-substituted RNA polymerases. This analog is a good substrate for Escherichia coli RNA polymerase and can be used to initiate the RNA chain. Incorporation of this analog into the total RNA synthesized was about 60% of that observed for ATP, independent of the templates used. Its Km values (22 and 118 microM) are twofold higher and its Vmax values (45 and 59 nmol/min/mg of enzyme) are 40% lower than those for ATP using calf thymus DNA and poly[d(A-T)], respectively, as template. For abortive initiation reaction using pAR1435 plasmid DNA as template, the Km and Vmax values of this analog are 2.7 times higher and 7 times lower, respectively, than those of ATP. With its desirable spectroscopic properties, (gamma-1,5-EDANS)ATP is a good probe for the studies of nucleotide-protein interactions, active site mapping of RNA polymerase, and other ATP-utilizing biological systems.

journal_name

Arch Biochem Biophys

journal_title

Archives of biochemistry and biophysics

authors

Wu FY,Abdulwajid AW,Solaiman D

doi

10.1016/0003-9861(86)90311-5

subject

Has Abstract

pub_date

1986-05-01 00:00:00

pages

564-71

issue

2

eissn

0003-9861

issn

1096-0384

pii

0003-9861(86)90311-5

journal_volume

246

pub_type

杂志文章

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