Identification and partial characterization of the xanthine oxidase transitions of the milk fat globule membrane.
Differential scanning calorimetry of bovine milk fat globule membranes (MFGM) yields five to eight transitions, depending on the conditions employed during isolation and assay of the membranes. Transitions A, B, and C were shown in a previous publication to derive from lipid melting, while transition D was found to stem from the unfolding of a structural protein termed butyrophilin [K. C. Appell, T. W. Kennan, and P. S. Low (1982) Biochim. Biophys. Acta 690, 243-250]. In this report we present evidence that the E1, E2, and F endotherms derive from the major MFGM protein, xanthine oxidase. Support for this contention derives from (i) thermal gel analysis; (ii) thermal inactivation analysis; (iii) comparison of the calorimetric properties of endotherms I, II, and III of purified xanthine oxidase with transitions E1, E2, and F of MFGM; (iv) comparison of the properties of a peculiar exotherm in scans of both the purified enzyme and MFGM; and (v) examination of the effects of specific ligands, reducing agents, and pH on both the xanthine oxidase and MFGM transition. The existence of three independent endotherms (I, II, and III) in purified xanthine oxidase demonstrates that the enzyme is composed of multiple independent domains. The interconversion of transitions I (E1) and II (E2) with a change in the redox conditions of the medium implies that these two transitions may be manifestations of the interconvertible dehydrogenase and oxidase forms of the enzyme, respectively. The relative independence of the I/II transitions from transition III further shows that only slight interaction between the major domains of xanthine oxidase exists.
D000818:Animals; D002152:Calorimetry, Differential Scanning; D002384:Catalysis; D002417:Cattle; D008563:Membrane Lipids / Q000378:metabolism*; D008565:Membrane Proteins / Q000378:metabolism*; D008892:Milk / Q000201:enzymology* / Q000378:metabolism; D010084:Oxidation-Reduction; D013816:Thermodynamics; D014969:Xanthine Oxidase / Q000378:metabolism*
D008563:Membrane Lipids; D008565:Membrane Proteins; D014969:Xanthine Oxidase