Linked Life Data

2440871

Source:http://linkedlifedata.com/resource/pubmed/id/2440871

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
22
pubmed:dateCreated
1987-9-17
pubmed:abstractText
New fluorescent derivatives of dinucleoside monophosphates, (5'-AmNS)UpA/ApU/GpU/CpA, with a fluorophore, 1-aminonaphthalene-5-sulfonic acid (AmNS), attached to the first nucleotide of the dinucleoside monophosphates via a 5'-secondary amine linkage were synthesized in good yield. The chemical structure of (5'-AmNS)ApU was proved by the phosphodiesterase digestion followed by Whatman No. 3MM paper chromatographic and spectroscopic analysis of the digested products. The ability of these analogs to be incorporated into the 5' terminus of RNA chain forming fluorescent oligonucleotides by Escherichia coli RNA polymerase was studied in the presence of a synthetic DNA template. The enzymatic reaction of (5'-AmNS)UpA and [3H]UTP in the presence of poly(dA-dT) yielded (5'-AmNS)UpAp[3H]U in greater than 30% yield with the Km values of 5 and 2.5 microM and Vmax values of 17 and 25 nmol/min/mg of enzyme for (5'-AmNS)UpA and UpA, respectively. The structure of this fluorescent trinucleotide was identified by RNase A digestion and paper chromatographic analysis of the digested products. (5'-AmNS)UpA or (5'-AmNS)ApU exhibits two absorption maxima around 270 and 340-350 nm and a fluorescent emission maximum at 445 nm when excited at 340 nm. These spectral characteristics permit their use as energy donors for the transfer of energy to the intrinsic cobalt of the cobalt-substituted RNA polymerases. Upon hydrolysis of the phosphodiester bond of these analogs by venom phosphodiesterase, the absorption at 340 and 270 nm increased by 5 and 20%, respectively, while their fluorescence at 445 nm was enhanced by 25%. Thus, these analogs can be used for studying the dynamics of initiation and elongation reactions catalyzed by DNA-dependent RNA polymerases by absorption and fluorescence spectroscopies.
pubmed:grant
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Aug
pubmed:issn
0021-9258
pubmed:author
pubmed:issnType
Print
pubmed:day
5
pubmed:volume
262
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
10684-8
pubmed:dateRevised
2007-11-14
pubmed:meshHeading
pubmed:year
1987
pubmed:articleTitle
Synthesis and characterization of fluorescent dinucleotide substrate for the DNA-dependent RNA polymerase from Escherichia coli.
pubmed:publicationType
Journal Article, Research Support, U.S. Gov't, P.H.S., Research Support, U.S. Gov't, Non-P.H.S.