[25] Capsular poly-γ-d-glutamate synthesis in Bacillus licheniformis

This chapter provides an overview of capsular poly-γ-D-glutamate synthesis in Bacillus licheniformis. A membranous polyglutamyl synthetase complex (PGSC) from Bacillus licheniformis 9945A catalyzes a sequence of membrane-associated enzymatic reactions in which L-glutamic acid is activated, racemized, and polymerized to form a poly(γ-D-glutamyl) capsule. The molecular mass of these surface polymers exceeds 106 daltons. The capsule is important in the pathogenicity of Bacillus anthracis because of its ability to influence host–cell interactions. The in vitro determination of the overall reaction catalyzed by the membranous PGSC is based on measuring the rate of incorporation of L-[U-14C]glutamic acid into the high-molecular-weight polymeric products (poly-γ-D-glutamic acid, PGA). These polymers are chromatographically and electrophoretically immobile. The activity of the L-glutamic acid activating enzyme is determined by measuring the rate of an ATP-dependent formation of [14C]glutamyl hydroxamate.