Characterization of Acrylamidase Isolated from a Newly Isolated Acrylamide-Utilizing Bacterium, Ralstonia eutropha AUM-01

A mesophilic bacterium capable of utilizing acrylamide was isolated, AUM-01, from soil collected from leaf litter at Picnic Point on the UW-Madison campus. In minimal medium with acrylamide as the sole carbon and nitrogen source, a batch culture of AUM-01 completely converted 28.0 mM acrylamide to acrylic acid in 8 h and reached a cell density of 0.3 (A(600)). Afterward all the acrylic acid was degraded by 20 h with the cell density increasing to 1.9 (A(600)). The acrylamide-utilizing bacterium was identified as Ralstonia eutropha based on morphological observations, the BiOLOG GN2 MicroPlate(TM) identification system for Gram-negative bacteria, and additional physiological tests. An acrylamidase that hydrolyzes acrylamide to acrylic acid was purified from the strain AUM-01. The molecular weight of the enzyme from AUM-01 was determined to be 38 kDa by SDS-PAGE. The enzyme had pH and temperature optima of 6.3 and 55A degrees C, and the influence of different metals and amino acids on the ability of the purified protein to transform acrylamide to acrylic acid was evaluated. The enzyme from AUM-01 was totally inhibited by ZnSO4 and AgNO3.