Sulfation of p-nitrophenyl-N-acetyl-beta-D-galactosaminide with a microsomal fraction from cultured chondrocytes
Chick embryo chondrocyte microsomes containing intact Golgi vesicles took up 3'-phosphoadenosine-5'-phospho(TVS)sulfate ((TVS)PAPS) in a time- and temperature-dependent, substrate-saturable manner. When (TVS)PAPS and p-nitrophenyl-N-acetyl-beta-D-galactosaminide (pNP-GalNAc) were added to the incubation in the absence of detergent, the microsomes catalyzed the transfer of sulfate from (TVS)PAPS to pNP-GalNAc to form pNP-GalNAc-6-TVSO4. The apparent Km values for PAPS in the uptake and the pNP-GalNAc sulfation reactions were 2 X 10(-7) and 2 X 10(-6) M, respectively. The sulfation of pNP-GalNAc by the microsomal preparation was inhibited by detergent. The microsomal fraction also catalyzed the transfer of sulfate from (TVS)PAPS to oligosaccharides prepared from chondroitin. However, in contrast to the sulfation of pNP-GalNAc, the rate of sulfation of these oligosaccharides was low in the absence of detergent and was markedly stimulated when detergent was added. Sulfation of pNP-GalNAc by the freeze-thawed microsomes was inhibited when the octasaccharide prepared from chondroitin was present in the reaction mixture. As the PAPS that had been internalized in the microsomal vesicles was consumed in the sulfation of pNP-GalNAc, more (TVS)PAPS was taken up and the sulfated pNP-GalNAc was released from the vesicles. These observations suggest that pNP-GalNAc may serve as a model membrane-permeable substrate for study of the 6-sulfo-transferase reaction involved in sulfation of chondroitin sulfate in intact Golgi vesicles.
- Research Organization:
- Univ. of Illinois, Urbana
- OSTI ID:
- 6182046
- Journal Information:
- J. Biol. Chem.; (United States), Journal Name: J. Biol. Chem.; (United States) Vol. 24; ISSN JBCHA
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
59 BASIC BIOLOGICAL SCIENCES
ADDITIVES
AMINES
ANIMAL TISSUES
ANIMALS
ATP
BETA DECAY RADIOISOTOPES
BETA-MINUS DECAY RADIOISOTOPES
BIOCHEMICAL REACTION KINETICS
BIRDS
BODY
CARBOHYDRATES
CARTILAGE
CELL CONSTITUENTS
CELL MEMBRANES
CHICKENS
CONNECTIVE TISSUE
DAYS LIVING RADIOISOTOPES
DETERGENTS
EMBRYOS
EMULSIFIERS
EVEN-ODD NUCLEI
FOWL
FREEZING
ISOTOPE APPLICATIONS
ISOTOPES
KINETICS
LIGHT NUCLEI
MEMBRANE TRANSPORT
MEMBRANES
MICROSOMES
NUCLEI
NUCLEOTIDES
OLIGOSACCHARIDES
ORGANIC COMPOUNDS
ORGANOIDS
OXYGEN COMPOUNDS
PERMEABILITY
RADIOISOTOPES
REACTION KINETICS
SACCHARIDES
SUBSTRATES
SULFATES
SULFUR 35
SULFUR COMPOUNDS
SULFUR ISOTOPES
SURFACTANTS
TEMPERATURE DEPENDENCE
THAWING
TIME DEPENDENCE
TISSUES
TRACER TECHNIQUES
VERTEBRATES
WETTING AGENTS