Purification,characterization and reconstitution of CMP-N-acetylneuraminate hydroxylase from mouse liver |
| |
Authors: | Petra Schneckenburger Lee Shaw Roland Schauer |
| |
Affiliation: | (1) Biochemisches Institut der Christian-Albrechts-Universität zu Kiel, Olshausenstr 40, D-24098 Kiel, FRG |
| |
Abstract: | CMP-N-acetylneuraminate hydroxylase was isolated from mouse liver high speed supernatant with a yield of 0.4% and an apparent 1000-fold purification. The enzyme is a monomeric protein with a molecular weight of 66 kDa, as determined by gel filtration and SDS-PAGE. The hydroxylase system was reconstituted with Triton X-100-solubilized mouse liver microsomes and purified soluble or microsomal forms of cytochrome b5 reductase and cytochrome b5. The systems were characterized in detail and kinetic parameters for each system were determined.Abbreviations Neu5Ac N-acetyl--d-neuraminic acid - Neu5Gc N-glycoloyl--d-neuraminic acid - CMP-Neu5Ac cytidine-5-monophospho-N-acetylneuraminic acid - CMP-Neu5Gc cytidine-5-monophospho-N-glycoloylneuraminic acid - TCA trichloroacetic acid - Chaps 3-[(3-cholamidopropyl)-dimethylammonio]-1-propanesulphonate - SOD superoxide dismutaseEnzymes: CMP-N-acetylneuraminate: NADH oxidoreductase (N-acetyl hydroxylating) (E.C. 1.14.13.45), CMP-Neu5Ac hydroxylase; NADH: cytochrome b5 oxidoreductase (E.C. 1.6.2.2), cytochrome b5 reductase; hydrogen peroxide: hydrogen peroxide oxidoreductase, catalase (E.C. 1.11.1.6); superoxide:superoxide oxidoreductase (E.C. 1.15.1.1), superoxide dismutase.This paper is dedicated to Professor Harry Schachter on the occasion of his 60th birthday. |
| |
Keywords: | sialic acid N-glycoloylneuraminic acid hydroxylase protein purification cytochrome b5 electron transfer enzyme system reconstitution |
本文献已被 SpringerLink 等数据库收录! |