Chemical apparatus and process disinfecting – deodorizing – preser – Control element responsive to a sensed operating condition – Sensed condition is operating time and control is operation...
Patent
1990-09-28
1992-08-11
Warden, Robert J.
Chemical apparatus and process disinfecting, deodorizing, preser
Control element responsive to a sensed operating condition
Sensed condition is operating time and control is operation...
422103, 422 681, 422 70, 436 89, G01N 3368
Patent
active
051376954
DESCRIPTION:
BRIEF SUMMARY
FIELD OF THE INVENTION
This invention relates generally to apparatus and for the performance of sequential chemical processes. More particularly, the invention relates to apparatus for the N-terminal and C-terminal sequencing of peptides.
DESCRIPTION OF THE PRIOR ART
The linear sequence of the amino acid units in proteins and peptides is of considerable interest as an aid to understanding their biological functions and ultimately synthesizing compounds performing the same functions. Although a variety of techniques have been used to determine the linear order of amino acids, probably the most successful is known in various forms as the Edman Process. The Edman sequential degradation processes involve three stages: coupling, cleavage and conversion. In the coupling stage, phenylisothiocyanate reacts with the N-terminal .beta. amino group of the peptide to form the phenylthiocarbamyl derivative. In the cleavage step, anhydrous acid is used to cleave the phenylthiocarbamyl derivative to form the anilinothiazolinone. After extraction of the thiazolinone, the residual peptide is ready for the next cycle of coupling and cleavage reactions. Aqueous acid is used to convert the thiazolinone to the phenylthiohydantoin which may be analyzed in an appropriate manner, such as by chromatography.
Practical automated N-terminal peptide sequencing dates from the 1967 introduction of the liquid phase spinning cup sequencer in which the reactions proceed in a thin liquid film formed on the inside wall of rotating reaction cells. See Edman, P., and Begg, G., A Protein Sequenator, European Journal of Biochemistry, 1:80-91 (1967). A focal problem associated with the spinning cup sequencer is sample loss, particularly of short peptides. An alternative solid phase degradation method entails passing reagents and solvents in an appropriate program through a column packed with porous material such as a macroporous polystyrene matrix or preferably porous glass beads to which a peptide is attached covalently or by adsorption. In another, known type of automatic sequencer the peptide to be degraded is covalently linked to a gel-type of solid phase support contained within a tubular reaction chamber. Both the reaction chamber and the tubing by which it is connected to the sequencer may be formed from polytetrafluoroethylene, e.g., "Teflon". See Laursen, R. A., A Solid-Phase Peptide Sequenator, European Journal of Biochemistry, 20:89-102 (1971) and Shively, "Methods of Protein Mischaracterization" Humana Press, Clifton, N.J. (1986), Chapter 9.
A sequencer that employs gas phase reagents instead of liquid phase reagents at critical points in the Edman degradation was proposed in 1981. See Hewick, R. M., Hunkapillar, M. W., Hood, L. E., Dreyer, W. J., A Gas-Liquid Solid Phase Peptide and Protein Sequenator, The Journal of Biological Chemistry, 256:7990-7997 (1981), U.S. Pat. No. 4,603,114 and Shively, supra, Chapter 8, Section 314, p. 229. This device includes a two-part glass cartridge assembly which houses a miniature continuous flow glass reaction chamber in which the peptide sample is presented as a dispersion in a thin film of a polymeric quaternary ammonium salt, such as polybrene, supported on a porous glass fiber disk. Means are provided for disconnecting the cartridge from its mounting base each time the sample is loaded.
A modification of the Hewick, et al. sequencer is described by Hawke, Harris and Shively in Analytical Biochemistry, 147:315-330 (1985), and Shively, supra, Chapter 7, page 210, et. seq. This modification replaces the glass reactor cartridge assembly of Hewick, et al. with an all Teflon cartridge of similar design, thus providing an all Teflon delivery and reaction system. The sample is presented within the reaction chamber on a trimethylsilyated glass fiber disk. Hawke, et al., noting that Teflon is "self-sealing", report lower background levels and increased yields deemed to be consequent from a better seal achieved in the all Teflon design as compared to the seal observed with the Hewick glass cartridge. S
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Calaycay Jimmy R.
Rusnak Miro
Shively John E.
City of Hope
Irons Edward S.
Trembley T. A.
Warden Robert J.
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