Method carboxy terminal protein or peptide sequencing

Details Method carboxy terminal protein or peptide sequencing Method carboxy terminal protein or peptide sequencing

1995-04-24

1997-06-24

Alexander, Lyle A.

Chemistry: analytical and immunological testing

Peptide, protein or amino acid

Amino acid or sequencing procedure

436 90, C07K 102, C07K 1107

Patent

active

056416857

ABSTRACT:
The present invention provides for an efficient and novel method for the C-terminal sequencing of proteins or peptides by use of acetyl chloride or phosphoryl chloride by reaction with a suitable isothiocyanate for derivitazation of the carboxy terminus to a thiohydantoin amino acid derivative, under acidic conditions. Cleavage of the derivatized thiohydantoin amino acid may occur by use of thiocyanic acid and acetic acid in water and also by the novel means using a buffer and a potassium or sodium thiocyanate or potassium or sodium dithionite reagent. The present invention also provides for an novel and efficient means for the C-terminal sequencing of proteins or peptides by a two or three step process which comprises first reacting the peptide or protein with an acid chloride reagent, such as acetyl chloride, or phosphoryl chloride. The stable protein carboxy terminal amino acid chloride is then reacted with an organic isothiocyanate, an organic salt thiocyanate, or a metal thiocyanate to yield a thiohydantoin amino acid derivative. This thiohydantoin amino acid derivative may be cleaved using either acidic or basic condition, preferably using the novel reagents disclosed above.

REFERENCES:
patent: 5041388 (1991-08-01), Boyd et al.
patent: 5185266 (1993-02-01), Boyd et al.
Inglis, Adam S., "Chemical Procedures for C-Terminal Sequencing of Peptides and Proteins," Analytical Biochemistry 195, pp. 183-196 (1991).
Stark, George R., "Sequential Degradation of Peptides from Their Carboxyl Terinini with Ammonium Thiocyanate and Acetic Anhydride," Biochemistry, vol. 7, No. 5, May 1968, pp. 1796-1807 (1968).

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