Mechanistic and stereochemical studies on the biosynthesis of neomycin

Al-Feel, Walid Mohammud Rashid (1983) Mechanistic and stereochemical studies on the biosynthesis of neomycin. University of Southampton, Doctoral Thesis.

Record type: Thesis (Doctoral)

Abstract

Mechanistic and stereochemical studies on the biosynthesis of neomycin. This thesis is concerned with the elucidation of the mechanism and stereochemistry of the formation of various subunits of neomycin B and C. Particular emphasis is laid on neosamine C and neosamine B subunits. For this purpose, D-glucose and D-glucosamine specifically labelled with 3H and 1sC were administered to 48 h old cultures of S. fradiae (ATCC 10745), which were allowed to grow for a further 12 h period. After harvesting, the antibiotic was isolated, either as a mixture of neomycin B plus C or the two compounds, neomycin B and C, were separated using ion-exchange chromatography. The isolated antibiotics were degraded chemically to furnish rings I, II and IV as neosaminol C, 2-deoxystreptamine and neosaminol B. Results from experiments involving the incorporation of D-[3H,10C]glucose or glucosamine into neomycin subunits showed that these precursors were incorporated intact and one of the tritium atoms present at C-6 of the parent precursor was eliminated during the formation of neosamine B and C_ On the other hand, 2-deoxystreptamine retained both the tritium atoms originally present at the precursors. It was concluded that the loss of one of thetritium atoms from C-6 of the precursors occurred during the formation of the aminomethyl group of neosamine B and C. From this latter observation, a mechanism for the formation of neosamine rings of the antibiotic was proposed. This mechanism assumed that the aminomethyl group of the neosamine is formed through the sequence -CH20H--r CH=O --,CH2-NH2 in which the C-6 hydroxyl group is first oxidised to an aldehyde and the latter then transaminated to furnish the product. The stereochemistry of the transamination processes in the formation of neosamine B and C was elucidated, using serine hydroxymethyltransferase, purified from the liver of rabbit. This enzyme catalyses the exchange of the Pro-S hydrogen atom of glycine with the protons ofthe medium. The incubation of serine hydroxymethyltransferase with [3H,I4CJ-glycine obtained from neosamine B or C resulted in a completeloss of tritium from Ca of glycine. These results suggest that the net transformation from D-glucose or D-glucosamine to neosamines proceeds stereospecifically with inversion of configuration at the C-6. Although the mechanism through which the C-5 of neosamine C is epimerized, producing the L-idose configuration at this centre could not be studied due to the supply of wrongly labelled samples of D- [5-'H] -glucose bydmersham. Two mechanisms for the epimerization are considered in this thesis.

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