Open AccessCMP‐ N ‐acetylneuraminic acid hydroxylase from mouse liver and pig submandibular glands
Author(s) -
SHAW Lee,
SCHNECKENBURGER Petra,
SCHLENZKA Wiebke,
CARLSEN Jens,
CHRISTIANSEN Kirsten,
JÜRGENSEN Dorthe,
SCHAUER Roland
Publication year - 1994
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.1432-1033.1994.tb18583.x
Subject(s) - cytochrome b5 , biochemistry , cytochrome , reductase , microsome , enzyme , cytochrome c , chemistry , cytochrome p450 reductase , biology , microbiology and biotechnology , mitochondrion , coenzyme q – cytochrome c reductase
In this report, the nature of the protein components involved in the functioning of cytidine‐5′‐monophosphate‐ N ‐acetylneuraminic acid (CMP‐Neu5 Ac) hydroxylase in high‐speed supernatants of mouse liver has been investigated. Fractionation and reconstitution experiments showed that this enzyme system consists of NADH–cytochrome b 5 reductase, cytochrome b 5 and a 56‐kDa terminal electron acceptor having the CMP‐Neu5 Ac hydroxylase activity. This enzyme system is extracted in a soluble protein fraction; however, the amphipathic, usually membrane‐associated, forms of cytochrome b 5 and the reductase were found to predominate and are presumably the forms which support the turnover of the hydroxylase in vivo. Although the majority of cellular cytochrome b 5 and cytochrome b 5 reductase is membrane‐bound, the addition of intact microsomes elicited no significant increase in the hydroxylase activity of supernatants. Detergent‐solubilised microsomes, however, potently activated the hydroxylase, probably due to the greater accessibility of the cytochrome b 5 . Accordingly, in reconstitution experiments, pure hydrophilic cytochrome b 5 interacts more effectively with the hydroxylase than isolated amphipathic cytochrome b 5 . Studies on the CMP‐Neu5Ac hydroxylase system in fractionated porcine submandibular glands and bovine liver suggest that the composition of this enzyme system is conserved in all mammals possessing sialoglycoconjugates containing N ‐glycoloylneuraminic acid.