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 (A₆₀₀)). Afterward all the acrylic acid was degraded by 20 h with the cell density increasing to 1.9 (A₆₀₀). The acrylamide-utilizing bacterium was identified as Ralstonia eutropha based on morphological observations, the BiOLOG GN2 MicroPlate™ 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 55°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.
Cha
Minseok
M
Department of Bacteriology, University of Wisconsin-Madison, Madison, WI 53706, USA. Min-Seok.Cha@pnl.gov
Chambliss
Glenn H
GH
D020106:Acrylamide / Q000378:metabolism*; D000179:Acrylates / Q000378:metabolism; D001426:Bacterial Proteins / Q000737:chemistry / Q000302:isolation & purification; D015373:Bacterial Typing Techniques; D002244:Carbon / Q000378:metabolism; D003470:Culture Media / Q000737:chemistry; D020118:Cupriavidus necator / Q000145:classification / Q000201:enzymology* / Q000302:isolation & purification / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D020536:Enzyme Activators / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000378:metabolism; D004795:Enzyme Stability; D006836:Hydro-Lyases / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D008970:Molecular Weight; D009584:Nitrogen / Q000378:metabolism; D012988:Soil Microbiology; D013696:Temperature
D000179:Acrylates; D001426:Bacterial Proteins; D003470:Culture Media; D020536:Enzyme Activators; D004791:Enzyme Inhibitors; D020106:Acrylamide; D002244:Carbon; D006836:Hydro-Lyases; C036658:acrylic acid; D009584:Nitrogen