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Characterization of the in vitro synthesized arabinan of mycobacterial cell walls
1Department of Microbiology, Colorado State University, Fort Collins 80523, USA.
Abstract:
Previous studies have shown that polymerized [14C]arabinan can be synthesized from polyprenylphosphate-[14C]arabinose by the particulate enzymes of Mycobacterium smegmatis [R.E. Lee, K. Mikusová, P.J. Brennan and G.S Besra (1995) J. Am. Chem. Soc. 117, 11829-11832]. In the present investigation, the [14C]arabinan product was biochemically characterized. Sizing chromatography revealed a molecular weight consistent with that expected from mature arabinan. Digestion of the [14C]arabinan with a mixture of arabinases produced oligo[14C]arabinoside fragments including hexa[14C]arabinoside and tetra[14C]arabinoside which originated from the non-reducing terminal regions of the polymer, and di[14C]arabinoside from the internal regions of the polymer. These arabinoside fragments represent the major known structural motifs that comprise the arabinan segment of arabinogalactan and lipoarabinomannan. The presence of [14C]arabinose in both the internal and external regions of the [14C]arabinan suggests that polyprenylphosphate-arabinose is the major, and perhaps the only, donor of arabinosyl residues in mycobacteria.
Insights
Mycobacterium smegmatis synthesizes [14C]arabinan using polyprenylphosphate-[14C]arabinose. Biochemical characterization confirmed its structure and arabinose donation to key mycobacterial components.
Area of Science:
- Microbiology
- Biochemistry
- Glycobiology
Background:
- Polymerized [14C]arabinan synthesis in Mycobacterium smegmatis was previously demonstrated.
- The arabinan polymer is a crucial component of mycobacterial cell wall structures like arabinogalactan and lipoarabinomannan.
Purpose of the Study:
- To biochemically characterize the synthesized [14C]arabinan.
- To elucidate the role of polyprenylphosphate-arabinose in arabinan synthesis.
Main Methods:
- Sizing chromatography to determine molecular weight.
- Enzymatic digestion with arabinases to generate oligosaccharide fragments.
- Analysis of fragment origin (terminal vs. internal regions).
Main Results:
- Sizing chromatography confirmed a molecular weight consistent with mature arabinan.
- Arabinase digestion yielded oligosaccharide fragments (hexa-, tetra-, and diarabinosides).
- Fragments originated from both non-reducing terminal and internal regions of the polymer.
Conclusions:
- Polyprenylphosphate-arabinose is the primary, possibly sole, arabinosyl residue donor in mycobacteria.
- The characterized [14C]arabinan fragments represent key structural motifs in mycobacterial cell walls.
- This study provides biochemical evidence for arabinan biosynthesis pathways in mycobacteria.