Extraction And Characterization Of Transferrin From Goose Serum

Goose blood is precious animal protein resource and contains many kind of proteins, such as transferrins, immunoglobulins, which have biological function. Until now, a little research has been done about goose plasma transferrin. The purpose of this thesis is the separation of main functional protein from goose blood, and the study on its physic-chemical and biological functional properties.After goose blood was centrifuged by refrigerated centrifuge, it was separated from blood, and then was frozen dried, and kept in low temperature. The content of globins was determined by cellulose acetate film electrophoresis, sephadex gel on G-100 and test of antibacterial ability. It was found that B-globulin was the major composition of goose plasma, the content of P-globulin was 34.809 mg/dl, and the content of r-globulin was found less part of goose plasma.The goose plasma transferrin was separated from goose blood by ion-exchange on DEAE-52 and sephadex gel on G-100. The purity of apo-TF was determined and found to be 98% by RP-HPLC. The molecular weight of Transferrin was about 77,820D by SDS-PAGE. By observation of dissolution of protein at different pH, the pI of transferrin was about 5.6. Fe was separated from transferrin by EDTA-acetic acid buffer, apo-transferrin obtained was white flock, which was dissolved in water easily. The denaturalization temperature of goose transferrrin was about 60C~90 C.Absorption of 2% (w/w) holo-Transferrin at 465nm was about 0.5. NaHCC>3, pH and temperature all influenced the ability of Transferrin combining Fe. When pH was 7.8, the ability of Transferrrin combining Fe was best, while pH was under 6.5, the ability of Transferrin combining Fe decreased remarkably. The addition of NaHCC>3 was availed for the ability of Transferrin combing Fe. When the concentration of NaHCO3 was beyond 100mmol/L, the effect wasn't remarkable and when Transferrin was kept under 70 C, the ability of Transferrin combining Fe decreased a little and the range wasn't beyond 3%. When tranferrin was deposed beyond 70C , the ability of Transferrin combining Fe decreased remarkably. So the temperature of sterilization under 70 C rebounded to keep the ability of Transferrin combining Fe. The influence of Fe3+ to molecule structure of holo-Transferrin and interactions of apo-and holo-transferrin with Al3+ were studied by circular dichroism (CD), UV-visible, and fluorescence spectrometry. The result of experiment showed that Transferrin can combine two Al3+, but Al3+ can't replace Fe3+.The influence of temperature, iron saturation and pH on antibacterial activity was studied. Transferrin restrained G+ better than G-. When iron saturation increased, the ability of Transferrin restraining bacterium decreased. Fe3+ maybe changed the molecule structure of Transferrin, when Transferrin combined Fe3+, which influenced the ability of Transferrin restraining bacterium. When pH was 7, the ability of Transferrin restraining bacterium was best. When pH was between 5~9,the change of pH influenced the ability of Transfrrin restraining G" a little, while the change of pHinfluenced the ability of Transferrrin restraining G+ greatly. When pH < 5.0 or pH > 9.0, the ability of Transferrin restraining bacterium was influenced greatly. When temperature of deposing transferrin was between 65C~70C, which influenced the ability of Transferrin restraining bacterium little. The increasing of concentration of cation will influence the ability of Transferrin restraining bacterium.