Abstract
A careful examination of two strains of Actinomyces viscosus, T14V (virulent) and T14AV (avirulent), revealed no qualitative or quantitative difference in 6-deoxyhexose content of their cell surface. For a further study of the role of these sugars in cell surface-related phenomena, the stereochemical configuration of deoxyhexoses of A. viscosus T14 was established by two complementary approaches. (i) Examination of the biosynthetic pathway was found to lead to the formation of both 6-deoxy-l-talose and 6-deoxy-l-mannose and showed no differences in the ability of either bacterial strain, A. viscosus T14V or T14AV, to produce the precursors of these cell wall components. The biosynthetic pathway for 6-deoxy-l-talose and 6-deoxy-l-mannose was found to originate from deoxy-thymidine diphosphate (dTDP)-d-glucose, which in turn is converted to dTDP-4-keto-6-deoxy-d-glucose. Epimerization at carbons 3 and 5 of the hexose moiety of dTDP-4-keto-6-deoxy-d-glucose is followed by stereospecific reduction with reduced nicotinamide adenine dinucleotide phosphate to yield dTDP-6-deoxy-l-talose and dTDP-6-deoxy-l-mannose. In cell-free extracts of both A. viscosus T14 and T14AV, an identical ratio of 6-deoxy-l-talose to 6-deoxy-l-mannose of 1:8 was produced. Known precursors for the d-isomers of the same 6-deoxyhexoses such as guanosine diphosphate-d-mannose and dTDP-d-mannose were not converted by A. viscosus T14 cell-free extracts. (ii) Isolation of 6-[U-(14)C]deoxytalose and 6-[U-(14)C]deoxymannose from both strains of A. viscosus T14 was carried out by growing cells in a medium containing d-[U-(14)C]glucose. Again no qualitative or quantitative difference was noticeable between the two strains when 6-deoxy-hexoses were released from whole cells or purified cell walls by acid hydrolysis. Radioactive 6-[U-(14)C]deoxytalose isolated from the cell surface was used in an isotope dilution experiment to establish the stereochemical configuration of this 6-deoxyhexose. The radioactive sugar was mixed with unlabeled standard d- or l-6-deoxyhexose, respectively, and conversion to the corresponding 1-phenylflavazole derivative was carried out. Recrystallization to constant specific activity identified the radioactive sugar isolated from A. viscosus to be the l-isomer. A facile synthesis of the rare sugars 6-deoxy-l-talose and 6-deoxy-d-talose is reported.