TY - JOUR
T1 - Detection of halogenated organic compounds using immobilized thermophilic dehalogenase
AU - Bachas-Daunert, Philip G.
AU - Sellers, Zachariah P.
AU - Wei, Yinan
PY - 2009/10
Y1 - 2009/10
N2 - Environmental pollutants containing halogenated organic compounds can cause a plethora of health problems. Detection, quantification, and eventual remediation of halogenated pollutants in the environment are important to human well-being. Toward this end, we previously identified a haloacid dehalogenase, L-HADST, from the thermophile Sulfolobus tokodaii. This thermophilic enzyme is extremely stable and catalyzes, stereospecifically, the dehalogenation of l-2-haloacids. In the current study, we covalently linked L-HADST to an N-hydroxysuccinimidyl Sepharose resin to construct a highly specific sensor with long shelf life for the detection of l-2-haloacids. The enzyme-modified resin was packed into disposable columns. Samples containing l-2-haloacids were first incubated in the column, and were then collected to quantify the chloride produced through the breakdown of the substrate. The optimum pH of the immobilized enzyme is around 9.5, similar to that of the soluble protein. Its catalytic activity increased with temperature up to the highest temperature measured (50 °C). The resin could be fully regenerated after multiple reaction cycles and retained 70% of the initial activity after being stored at 4 °C for 6 months. The L-HADST-modified resin could be used to breakdown and quantify l-2-haloacids spiked in the simulated environmental samples, indicating dehalogenases from extremophiles can potentially be employed in the detection and decontamination of l-2-haloacids.
AB - Environmental pollutants containing halogenated organic compounds can cause a plethora of health problems. Detection, quantification, and eventual remediation of halogenated pollutants in the environment are important to human well-being. Toward this end, we previously identified a haloacid dehalogenase, L-HADST, from the thermophile Sulfolobus tokodaii. This thermophilic enzyme is extremely stable and catalyzes, stereospecifically, the dehalogenation of l-2-haloacids. In the current study, we covalently linked L-HADST to an N-hydroxysuccinimidyl Sepharose resin to construct a highly specific sensor with long shelf life for the detection of l-2-haloacids. The enzyme-modified resin was packed into disposable columns. Samples containing l-2-haloacids were first incubated in the column, and were then collected to quantify the chloride produced through the breakdown of the substrate. The optimum pH of the immobilized enzyme is around 9.5, similar to that of the soluble protein. Its catalytic activity increased with temperature up to the highest temperature measured (50 °C). The resin could be fully regenerated after multiple reaction cycles and retained 70% of the initial activity after being stored at 4 °C for 6 months. The L-HADST-modified resin could be used to breakdown and quantify l-2-haloacids spiked in the simulated environmental samples, indicating dehalogenases from extremophiles can potentially be employed in the detection and decontamination of l-2-haloacids.
KW - Dehalogenase
KW - Electrochemistry
KW - Enzyme immobilization
KW - Thermophilic
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U2 - 10.1007/s00216-009-3057-5
DO - 10.1007/s00216-009-3057-5
M3 - Article
C2 - 19714319
AN - SCOPUS:70349777273
SN - 1618-2642
VL - 395
SP - 1173
EP - 1178
JO - Analytical and Bioanalytical Chemistry
JF - Analytical and Bioanalytical Chemistry
IS - 4
ER -