Cellulose Microfibril Formation by Surface-Tethered Cellulose Synthase Enzymes.

Cellulose microfibrils are pseudo crystalline arrays of cellulose chains that are synthesized by cellulose synthases. The enzymes are organized into large membrane-embedded complexes in which each enzyme likely synthesizes and secretes a β-(1→4) glucan. The relationship between the organization of the enzymes in these complexes and cellulose crystallization has not been explored. To better understand this relationship, we used atomic force microscopy to visualize cellulose microfibril formation from nickel film-immobilized bacterial cellulose synthase enzymes (BcsA-Bs), which in solution only form amorphous cellulose from monomeric BcsA-B monomeric complexes. Fourier Transform Infrared spectroscopy and X-ray diffraction techniques show that surface tethered BcsA-Bs synthesize highly crystalline cellulose II in the presence of UDP-Glc, the allosteric activator cyclic-di-GMP, as well as magnesium. The cellulose II cross-section/diameter as well as crystal size and crystallinity depend on the surface density of tethered enzymes as well as the overall concentration of substrates. Our results provide the correlation between cellulose microfibril formation and the spatial organization of cellulose synthases.