Enhancement of the specificity of an enzyme-based biosensor for L-tryptophan.
A new selective amperometric biosensor for reagentless L-tryptophan determination has been developed using immobilized tryptophan-2-monooxygenase (TMO, EC 1.13.12.3). This enzyme-based biosensor provides a rapid-response detection system for concentrations of L-tryptophan between 25 and 1,000 microM in a batch mode system and between 100 and 50,000 microM in a flow-injection mode. The response time was 30 seconds, and the total analysis time was less than 3 minutes. The biosensor retained catalytic activity and fidelity of phenylalanine and tryptophan response for greater than 4 months with repeated usage. The biosensor selectivity to L-tryptophan was dramatically increased relative to phenylalanine when a competitive inhibitor of TMO, indole acetamide (IA), was included. The biosensor was successfully used for L-tryptophan determination in nutrition broth, giving values identical to those determined by HPLC analysis.
Simonian
A L
AL
Texas A&M University, Department of Biochemistry and Biophysics, College Station 77843-2128, USA. als@pop.tamu.edu
Rainina
E I
EI
Fitzpatrick
P F
PF
Wild
J
J
D015374:Biosensing Techniques; D004563:Electrochemistry; D004800:Enzymes, Immobilized; D004926:Escherichia coli / Q000201:enzymology; D007700:Kinetics; D010649:Phenylalanine / Q000378:metabolism; D011549:Pseudomonas / Q000201:enzymology; D011994:Recombinant Proteins; D012680:Sensitivity and Specificity; D014364:Tryptophan / Q000032:analysis* / Q000378:metabolism; D014365:Tryptophan Hydroxylase
D004800:Enzymes, Immobilized; D011994:Recombinant Proteins; D010649:Phenylalanine; D014364:Tryptophan; D014365:Tryptophan Hydroxylase