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Protein sequence analysis using Hewlett-Packard biphasic sequencing cartridges in an applied biosystems 473A protein
S Tang1, J Mozdzanowski, K R Anumula
1Bioanalytical Sciences, Research & Development, SmithKline Beecham Pharmaceuticals, King of Prussia, Pennsylvania 19406, USA.
Journal of Protein Chemistry
|March 11, 1999
Summary
A new Hewlett-Packard cartridge enables easier protein sequencing of difficult samples, like formulated drugs, using the Applied Biosystems sequencer. This method provides sequence data sufficient for protein identification and DNA probe preparation.
Area of Science:
- Biochemistry
- Analytical Chemistry
- Molecular Biology
Background:
- Edman degradation is a standard method for protein sequencing.
- Analyzing complex protein samples, such as formulated drugs, presents challenges in traditional sequencing.
- Hewlett-Packard developed an adsorptive biphasic sequencing cartridge to address these challenges.
Purpose of the Study:
- To demonstrate the utility of the Hewlett-Packard adsorptive biphasic sequencing cartridge with the Applied Biosystems 473A protein sequencer.
- To evaluate the compatibility of the cartridge with standard sequencer solvents and reagents.
- To assess the effectiveness of this system for analyzing challenging protein samples.
Main Methods:
- Utilized an adsorptive biphasic sequencing cartridge (Hewlett-Packard) in an Applied Biosystems 473A protein sequencer.
- Analyzed protein samples containing common excipients, salts, and detergents.
- Performed simple modifications to the Applied Biosystems sequencer to accommodate the cartridge.
Main Results:
- Successfully demonstrated protein sequence analysis using the combined system.
- Confirmed compatibility of Applied Biosystems sequencer solvents and reagents with the Hewlett-Packard cartridge.
- Generated sequence information up to ten residues for complex samples without optimization.
- Identified proteins and prepared DNA probes for novel protein cloning.
Conclusions:
- The adsorptive biphasic sequencing cartridge extends the utility of the Applied Biosystems 473A sequencer for analyzing difficult protein samples.
- This method provides sufficient sequence data for protein identification and molecular cloning applications.
- The system offers a practical approach for sequencing formulated protein drugs and other complex biological samples.
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