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Decay accelerating factor (DAF) is anchored to membranes by a C-terminal glycolipid. [Decay accelerating factor]

Conference · · Fed. Proc., Fed. Am. Soc. Exp. Biol.; (United States)
OSTI ID:5453544
; ; ;

Purified 70 kDa membrane (m) DAF incorporates into cells when added in vitro. A 2 kDa smaller DAF form which functions extrinsically like C4bp but is unable to incorporate can be isolated from urine (u). Because of common deficits of mDAF and acetylcholinesterase (AChE) in erythrocytes (E) of patients with paroxysmal nocturnal hemoglobinuria (PNH), mDAF was analyzed for a O-terminal glycolipid membrane anchor similar to that in E AChE. Incubation of E with phosphatidylinositol-specific phospholipase C, an enzyme which cleaves a similar glycolipid anchor in trypanosome variant surface glycoproteins (mfVSGs), released 20% of the DAF antigen. The released DAF species resembled uDAF in size, extrinsic model of C4b2a decay, and lack of hydrophobicity. Reductive radiomethylation of mDAF with (/sup 14/C)HCHO and NaCNBH/sub 3/ revealed ethanolamine and glucosamine in proportions similar to those in the E AChE glycolipid anchor. Papain cleavage of radiomethylated mDAF released the labeled ethanolamine and glucosamine in small O-terminal fragments from the residual DAF that retained N-terminal Asp. Following labeling of the anchors of mDAF and E AChE with the lipophilic photoreagent 3-trifluoromethyl-3-(m-(/sup 125/I)iodophenyl)diazirine, cleavage at the glucosamine residue by deamination quantitatively released the label from both proteins. Biosynthetic labeling of Hela cells with (/sup 3/H)ethanolamine resulted in rapid /sup 3/H incorporation into both 48 kDa proDAF and 70 kDa mDAF. These data indicate that mDAF is anchored by a glycolipid similar to that in E AChE, mfVSGs and Thy-1 antigen and raise the possibility that a defect in the assembly or attachment of this structure could account for the deficits of mDAF and E AChE in PNH.

Research Organization:
Case Western Reserve Univ., Cleveland, OH
OSTI ID:
5453544
Report Number(s):
CONF-8604222-
Journal Information:
Fed. Proc., Fed. Am. Soc. Exp. Biol.; (United States), Journal Name: Fed. Proc., Fed. Am. Soc. Exp. Biol.; (United States) Vol. 45:3; ISSN FEPRA
Country of Publication:
United States
Language:
English

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Related Subjects

550201* -- Biochemistry-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
ALDEHYDES
AMINES
ANTIGENS
BETA DECAY RADIOISOTOPES
BIOCHEMICAL REACTION KINETICS
BIOCHEMISTRY
BIOLOGICAL MATERIALS
BLOOD
BLOOD CELLS
BODY FLUIDS
CARBOHYDRATES
CARBON 14 COMPOUNDS
CARBOXYLESTERASES
CELL CONSTITUENTS
CELL DIFFERENTIATION
CELL MEMBRANES
CHEMISTRY
CHOLINESTERASE
DAYS LIVING RADIOISOTOPES
ELECTRON CAPTURE RADIOISOTOPES
ENZYMES
ERYTHROCYTES
ESTERASES
FORMALDEHYDE
GLUCOPROTEINS
GLUCOSAMINE
HELA CELLS
HEXOSAMINES
HEXOSES
HORMONES
HYDROLASES
INTERMEDIATE MASS NUCLEI
IODINE 125
IODINE ISOTOPES
ISOTOPE APPLICATIONS
ISOTOPES
KINETICS
LABELLED COMPOUNDS
LIPASE
MATERIALS
MEMBRANE PROTEINS
MEMBRANES
MONOSACCHARIDES
NUCLEI
ODD-EVEN NUCLEI
ORGANIC COMPOUNDS
PROTEINS
RADIOISOTOPES
REACTION KINETICS
RECEPTORS
SACCHARIDES
TRACER TECHNIQUES
TRITIUM COMPOUNDS