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Luminescent terbium protein labels for time-resolved microscopy and screening
Harsha E Rajapakse1, D Rajasekhar Reddy, Shabnam Mohandessi
1Department of Chemistry, University of Illinois at Chicago, 845 W. Taylor St., Chicago, IL 60607, USA.
Angewandte Chemie (International Ed. in English)
|June 4, 2009
Summary
Researchers developed novel terbium chelate conjugates that bind bacterial dihydrofolate reductase fusion proteins. This terbium luminescence allows for sensitive, time-resolved detection of proteins in cells.
Area of Science:
- Bioconjugation Chemistry
- Biochemistry
- Analytical Chemistry
Background:
- Dihydrofolate reductase (DHFR) is a crucial enzyme in bacterial folate metabolism, making it a target for antibiotics like trimethoprim.
- Developing sensitive detection methods for specific proteins is vital in biological research and diagnostics.
Purpose of the Study:
- To synthesize and characterize novel heterodimeric conjugates linking trimethoprim to terbium chelates.
- To evaluate the binding affinity of these conjugates to Escherichia coli dihydrofolate reductase (EcDHFR) fusion proteins.
- To demonstrate the utility of terbium luminescence for sensitive protein detection.
Main Methods:
- Covalent linkage of trimethoprim to sensitized terbium chelates to form heterodimeric conjugates.
- Affinity determination using techniques to measure binding to EcDHFR fusion proteins.
- Utilizing terbium luminescence for time-resolved detection of labeled proteins in vitro and in live mammalian cells.
Main Results:
- The synthesized conjugates exhibited nanomolar binding affinity for EcDHFR fusion proteins.
- Terbium luminescence provided a sensitive and time-resolved signal for protein detection.
- Successful detection of labeled proteins was achieved both in vitro and on the surface of living mammalian cells.
Conclusions:
- Heterodimeric conjugates of trimethoprim and terbium chelates offer a promising tool for specific protein detection.
- Terbium luminescence provides a highly sensitive and time-resolved detection modality for biological applications.
- This approach has potential applications in bacterial enzyme studies and cellular imaging.
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