The use of radioactive cysteine methyl ester for labeling glycosylated molecules oxidized by periodate or neuraminidase plus galactose oxidase

Journal Article · Thu Mar 01 00:00:00 EST 1984 · Arch. Biochem. Biophys.; (United States)

DOI:https://doi.org/10.1016/0003-9861(84)90186-3· OSTI ID:7135221

Mitchell, R N; Harrison, E H; Bowers, W E

Treatment of rat lymph node cells with periodate or neuraminidase plus galactose oxidase initiates blast transformation and cell division of T lymphocytes. Either treatment introduces aldehyde functions onto glycosylated molecules of the plasma membrane. Reduction of the aldehydes with borohydride leads to a concentration-dependent inhibition of the mitogenic response. Cysteine methyl ester (Cys(Me), which can form a stable thiazolidine adduct with aldehydes, also inhibits mitogenesis in a concentration-dependent manner. Maximum inhibition is achieved at Cys(Me) concentrations about 10-fold lower than those required for borohydride (0.4 and 5 mM, respectively). (35S)Cys(Me) has been synthesized and compared with (/sup 3/H)borohydride as a labeling reagent for molecules on the plasma membrane oxidized by periodate or neuraminidase plus galactose oxidase. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of labeled whole cell lysates or of crude membrane fractions prepared from labeled cells revealed that the same oxidized molecules are specifically labeled with both reagents. Homogenates of cells labeled with either radioactive reagent were fractionated by differential and isopycnic centrifugation. The fractions were analyzed for radioactivity and for a number of marker constituents localized in various subcellular organelles. Following treatment with either reagent, the radioactivity that was covalently incorporated into macromolecules was primarily associated with sedimentable components that distributed among the fractions like plasma membrane markers. When compared with (/sup 3/H)borohydride, Cys(Me) offers several advantages as a surface labeling reagent for glycosylated plasma membrane molecules, chiefly the possibility of preparing reagents labeled with isotopes other than tritium, including those like /sup 35/S, which are much stronger radioactive emitters.

• Research Organization: Rockefeller University, New York, New York
• OSTI ID: 7135221
• Journal Information: Arch. Biochem. Biophys.; (United States), Vol. 229:2
• Country of Publication: United States
• Language: English

Related Subjects

59 BASIC BIOLOGICAL SCIENCES
CELL MEMBRANES
LABELLING
ALDEHYDES
CYSTEINE
ESTERS
LABELLED COMPOUNDS
LYMPHOCYTES
PERIODATES
RATS
SULFUR 35
TRITIUM COMPOUNDS
AMINO ACIDS
ANIMAL CELLS
ANIMALS
BETA DECAY RADIOISOTOPES
BETA-MINUS DECAY RADIOISOTOPES
BIOLOGICAL MATERIALS
BLOOD
BLOOD CELLS
BODY FLUIDS
CARBOXYLIC ACIDS
CELL CONSTITUENTS
CONNECTIVE TISSUE CELLS
DAYS LIVING RADIOISOTOPES
EVEN-ODD NUCLEI
HALOGEN COMPOUNDS
IODINE COMPOUNDS
ISOTOPES
LEUKOCYTES
LIGHT NUCLEI
MAMMALS
MATERIALS
MEMBRANES
NUCLEI
ORGANIC ACIDS
ORGANIC COMPOUNDS
ORGANIC SULFUR COMPOUNDS
OXYGEN COMPOUNDS
RADIOISOTOPES
RODENTS
SOMATIC CELLS
SULFUR ISOTOPES
THIOLS
VERTEBRATES
550201* - Biochemistry- Tracer Techniques