PMID: 6108211Sep 1, 1980Paper

Affinity chromatography on immobilised nucleotides. The synthesis, specificity and applications of immobilised inosine 5'-monophosphate

European Journal of Biochemistry
Y D Clonis, C R Lowe

Abstract

The synthesis and characterisation of two IMP analogues, 8-(6-aminohexyl)-ionosine 5'-monophosphate, Ahx8IMP, and inosine 2',3'-O-[1-(6-aminohexyl)-levulinic acid amide]-acetyl 5'-monophosphate, (AhxLvn)2',3'IMP, is described. These analogues were attached to CNBr-activated agarose through the terminal amino group of the spacer molecule. The immobilised-IMP analogues displayed specificity for the inosine-nucleotide-dependent enzyme, IMP dehydrogenase (IMP:NAD+ oxidoreductase, EC 1.2.1.14) but not for the NAD+-dependent enzymes, L-alanine and L-acetate dehydrogenases. Escherichia coli IMP dehydrogenase could be eluted biospecifically from immobilised 8-substituted and ribose-substituted IMP adsorbents with IMP, XMP and GMP. Multiple peaks of enzyme activity in the elution profiles were interpreted in terms of aggregation of the enzyme. A protocol for the large-scale purification of E. coli IMP dehydrogenase is proposed. Homogeneous enzyme of specific activity 9.1 units/mg was obtained in 50% overall yield, representing 14 mg pure protein from a 20-1 culture of E. coli. The two IMP analogues were inactive as substrates in the IMP dehydrogenase reaction.

References

Jan 1, 1976·Methods in Enzymology·K MosbachC Lowe
May 1, 1978·Archives of Biochemistry and Biophysics·P E BrodeliusN O Kaplan
Apr 28, 1973·Biochimica Et Biophysica Acta·R LamedM Wilchek
Sep 1, 1973·Journal of Bacteriology·P R Lambden, W T Drabble
Jun 1, 1974·The Biochemical Journal·I P TrayerR C Bottomley
Jun 1, 1974·Pharmacological Research Communications·G MichalG Weimann
Feb 1, 1969·Chemical & Pharmaceutical Bulletin·M Ikehara, S Uesugi
May 20, 1971·FEBS Letters·C R. Lowe, P D.G. Dean

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Citations

Aug 15, 1994·Journal of Biotechnology·N Labrou, Y D Clonis
May 7, 2014·Rapid Communications in Mass Spectrometry : RCM·Tanja C W Moerdijk-PoortvlietHenricus T S Boschker
Oct 26, 2005·Journal of Chromatography. a·Yannis D Clonis

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