The role of GumE in polymerization of the Xanthomonas campestris exopolysaccharide xanthan.

GALVAN, ESTELA M.; KAMPEL, MATÍAS; IELPI, LUIS

Puerto Madryn, Argentina

Congreso; XLV Reunión Anual de la Sociedad Argentina de Investigación en Bioquímica y Biología Molecular; 2010

Sociedad Argentina de Investigación en Bioquímica y Biología Molecular

Xanthan is a linear cellulosic b(1,4)-D-glucose polymer with trisaccharide side chains. Normal xanthan produced by wild type Xanthomonas campestris has an average molecular length of 0.9 mm (around 2,500 pentasaccharide repeat units). Biosynthesis and assembly of xanthan requires the enzymes GumB to GumM. We have already shown that gumB, gumC, and gumE mutant strains did not produce xanthan, but synthesized the lipid-linked repeat units. Longer xanthan molecules were obtained by overexpression of GumB and GumC. This increase in molecular length correlated with the increased viscosity observed. Here we studied GumE, a 432 amino acid protein. Hydropathy plotting of GumE showed a high degree of hydrophobicity, with ten transmembrane segments and two major hydrophilic loops, similar to other Wzy polymerases. A 507-bp deletion within gumE, removing the two hydrophilic loops and the three transmembrane segments between them, abolished xanthan biosynthesis. The full-length gumE expressed in trans fully restored xanthan production of gumE mutant strain. Overexpression of GumE in the wild-type background caused a significant decrease in xanthan viscosity (around 40-60 %) without changing the total amount of polymer produced. This effect was attributed to a shortening of the polymer chains. These results suggest that the levels of GumE, GumB, and GumC modulate the chain-length of xanthan.