Assembly and deacetylation of N-acetylglucosaminyl-plasmanylinositol in normal and affected paroxysmal nocturnal hemoglobinuria cells

Journal Article · Wed May 01 04:00:00 EDT 1991 · Proceedings of the National Academy of Sciences of the United States of America; (United States)

DOI:https://doi.org/10.1073/pnas.88.9.3762· OSTI ID:6056965

Hirose, Shinichi; Ravi, Lakshmeswari; Medof, M D ^[1]; Hazra, S V ^[2]

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1. Case Western Univ., Cleveland, OH (United States)
2. Akron General Medical Center, OH (United States)

Decay-accelerating factor (DAF) is anchored in cell membranes by a glycosyl-plasmanylinositol (GPI) moiety that is transferred to it en bloc in the rough endoplasmic reticulum. To analyze the biochemical reactions involved in preassembly of this structure, a human hematopoietic cell-free system was employed. Incubation of cell extracts with UDP-({sup 3}H)GlcNAc and butanol partitioning of reaction mixtures yielded two products similar in TLC mobility to intermediates described in Trypanoxoma brucei. Both species were sensitive to Bacillus thuringiensis phosphatidylinositol-specific phospholipase C, indicative of association of ({sup 3}H)GlcNAc label with a plasmanylinositol-containing acceptor. Kinetic and pulse-chase experiments indicated that the slower-migrating species was a product of the faster and that it, but not the faster, was sensitive to both GPI-specific phospholipase D and nitrous acid deamination, consistent with conversion of GlcNAc- to GlcN-plasmanylinositol. Lysates of normal and of affected blood leukocytes from two paroxysmal nocturnal hemoglobinuria (PNH) patients supported assembly of the two intermediates within 1 min. Thus, the initial enzymes mediating human GPI-anchor assembly are GlcNAc-plasmanylinositol transferase and GlcNAc-plasmanylinositol deacetylase, their substrates contain plasmanylinositols, and the products of their activities are normal in affected PNH cells.

• OSTI ID: 6056965
• Journal Information: Proceedings of the National Academy of Sciences of the United States of America; (United States), Journal Name: Proceedings of the National Academy of Sciences of the United States of America; (United States) Vol. 88:9; ISSN PNASA; ISSN 0027-8424
• Country of Publication: United States
• Language: English

Related Subjects

550201* -- Biochemistry-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
ALCOHOLS
ANIMAL CELLS
ANIMALS
BIOCHEMISTRY
BODY
BONE MARROW CELLS
BUTANOLS
CARBOHYDRATES
CELL CONSTITUENTS
CELL MEMBRANES
CHEMISTRY
CHROMATOGRAPHY
CONNECTIVE TISSUE CELLS
ENDOPLASMIC RETICULUM
ENZYMES
GLYCOLIPIDS
HEMATOPOIETIC SYSTEM
HYDROXY COMPOUNDS
IN VITRO
INOSITOLS
INVERTEBRATES
LIPIDS
MASTIGOPHORA
MEMBRANE PROTEINS
MEMBRANES
MICROORGANISMS
MONOSACCHARIDES
ORGANIC COMPOUNDS
PARASITES
PROTEINS
PROTOZOA
SACCHARIDES
SEPARATION PROCESSES
SOMATIC CELLS
THIN-LAYER CHROMATOGRAPHY
TRANSFERASES
TRYPANOSOMA