Isolation and characterization of ornitho-kininogen

Area of Science:

• Biochemistry
• Proteomics
• Comparative Physiology

Background:

• Kininogens are precursors to kinins, vasoactive peptides involved in various physiological processes.
• Mammalian kininogens exist in high and low molecular weight forms, with distinct structures and functions.
• The presence and characteristics of kininogens in avian species remain largely unexplored.

Purpose of the Study:

• To purify and characterize ornitho-kininogen from chicken blood plasma.
• To identify and determine the structure of the kinin liberated from ornitho-kininogen.
• To investigate the biological activity and specificity of the isolated chicken kinin.

Main Methods:

• Two-stage purification using chromatography (S-alkylated papain-Cellulofine and DEAE-5PW).
• Analysis by SDS-PAGE and disc/PAGE electrophoresis.
• Molecular mass determination using Ferguson plot analysis on SDS-PAGE.
• Peptide isolation via reversed-phase HPLC and primary structure determination.
• Assessment of biological activity on chicken smooth muscle and rat uterus.

Main Results:

• Ornitho-kininogen was successfully purified with a yield of 1.7 mg from 44 ml plasma.
• The purified protein exhibited a single band on SDS-PAGE and had a molecular mass of 74,000 Da.
• Chicken plasma contains high-Mr kininogen-like protein but lacks low-Mr kininogen.
• Bovine plasma kallikrein degraded ornitho-kininogen, releasing a kinin named ornitho-kinin (Arg-Pro-Pro-Gly-Phe-Thr-Pro-Leu-Arg).
• Ornitho-kinin induced chicken smooth muscle contraction and hypotension but not rat uterus contraction.

Conclusions:

• Chicken plasma contains a high molecular weight kininogen analogous to mammalian counterparts.
• A novel kinin, ornitho-kinin, was identified with structural similarities and functional differences compared to bradykinin.
• Ornitho-kinin exhibits species-specific biological activity, suggesting evolutionary adaptations in the kinin system.