The assembly of alpha-glucan by GlgE and GlgB in mycobacteria and streptomycetes.

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Actinomycetes, such as mycobacteria and streptomycetes, synthesize a-glucan with a-1,4 linkages and a-1,6 branching to help evade immune responses and to store carbon. a-Glucan is thought to resemble glycogen except for having shorter constituent linear chains. However, the fine structure of a-glucan and how it can be defined by the maltosyl transferase GlgE and branching enzyme GlgB were not known. Using a combination of enzymolysis and mass spectrometry, we compared the properties of a-glucan isolated from actinomycetes with polymer synthesized in vitro by GlgE and GlgB. We now propose the following assembly mechanism. Polymer synthesis starts with GlgE and its donor substrate, a-maltose 1-phosphate, yielding a linear oligomer with a degree of polymerization (~16) sufficient for GlgB to introduce a branch. Branching involves strictly intra-chain transfer to generate a C chain (the only constituent chain to retain its reducing end), which now bears an A chain (a non-reducing end terminal branch that does not itself bear a branch). GlgE preferentially extends A chains allowing GlgB to act iteratively to generate new A chains emanating from B chains (non-terminal branches that themselves bear a branch). Although extension and branching occurs primarily with A chains, the other chain types are sometimes extended and branched such that some B chains (and possibly C chains) bear more than one branch. This occurs less frequently in a-glucans than in classical glycogens. The very similar properties of cytosolic and capsular a-glucans from M. tuberculosis implies GlgE and GlgB are sufficient to synthesize them both.