Biosynthetic Studies on Geldanamycin and Pactamycin
Potgieter, Maria
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https://hdl.handle.net/2142/70248
Description
Title
Biosynthetic Studies on Geldanamycin and Pactamycin
Author(s)
Potgieter, Maria
Issue Date
1983
Department of Study
Chemistry
Discipline
Chemistry
Degree Granting Institution
University of Illinois at Urbana-Champaign
Degree Name
Ph.D.
Degree Level
Dissertation
Keyword(s)
Chemistry, Organic
Abstract
The formation of the m-C(,7)N units of geldanamycin and pactamycin via the shikimate pathway was studied. {('13)C(,6)}glucose was used to determine the incorporation of intact carbon units into both molecules. This revealed that the respective m-C(,7)N units were derived from three-carbon and four-carbon subunits (specifically phosphoenol pyruvate and erythrose-4-phosphate). In the case of geldanamycin C-15, C-16 and C-21 originated from the C(,3) unit and C-17 through C-20 from the C(,4) unit. This pattern was obtained by analysis of coupling patterns and coupling constants in the ('13)C NMR spectrum as well as homonuclear ('13)C-('13)C decoupling. Similarly, in the case of pactamycin C-2'' through C-5'' were derived from the C(,4) unit and C-1'', C-6'' and C-7'' from the C(,3) unit. The ('13)C NMR spectra of geldanamycin and pactamycin labeled by {('13)C(,6)}-glucose were analysed in terms of the incorporation of intact subunits into the rest of the molecules.
Radiolabeled glucose, 3-dehydroquinic, 3-dehydroshikimic and shikimic acids were used to study the role of the early shikimate pathway in m-C(,7)N biosynthesis. The cells of the geldanamycin-producing organism were equally permeable for all four precursors, while cell penetration by shikimic acid (SA) was markedly lower for the pactamycin producer. The branch point for geldanamycin is probably at SA and for pactamycin most likely either SA or 3-dehydroshikimate. 3-Amino-5-hydroxybenzoic acid was determined to be the advanced precursor for the m-C(,7)N unit of geldanamycin but it was not incorporated into pactamycin. Conversely, m-aminobenzoic acid, the seven-carbon precursor for pactamycin, was not incorporated into geldanamycin.
Hydrogen retention during N- and C-methylation of the pactamycins was studied using L-{methyl-('13)C('2)H(,3)}methionine. Only 7-deoxypactamycin and 7-deoxyisopactamycate were isolated from these fermentations, indicating that a kinetic isotope effect existed in the oxidation of C-7 of pactamycin. A study of fast atom bombardment mass spectra and ('13)C and ('2)H NMR spectra revealed that N- and C-methylation occurred with complete retention of the S-methyl hydrogen atoms.
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