Immunochemical characterization of the brain glutamate binding protein

Thesis/Dissertation · Wed Jan 01 04:00:00 EST 1986

OSTI ID:6994783

Roy, S

A glutamate binding protein (GBP) was purified from bovine and rat brain to near homogeneity. Polyclonal antibodies were raised against this protein. An enzyme-linked-immunosorbent-assay was used to quantify and determine the specificity of the antibody response. The antibodies were shown to strongly react with bovine brain GBP and the analogous protein from rat brain. The antibodies did not show any crossreactivity with the glutamate metabolizing enzymes, glutamate dehydrogenase, glutamine synthetase and glutamyl transpeptidase, however it crossreacted moderately with glutamate decarboxylase. The antibodies were also used to define the possible physiologic activity of GBP in synaptic membranes. The antibodies were shown: (i) to inhibit the excitatory amino-acid stimulation of thiocyanate (SCN)flux, (ii) had no effect on transport of L-Glutamic acid across the synaptic membrane, and (iii) had no effect on the depolarization-induced release of L-glutamate. When the anti-GBP antibodies were used to localize and quantify the GBP distribution in various subcellular fractions and in brain tissue samples, it was found that the hippocampus had the highest immunoreactivity followed by the cerebral cortex, cerebellar cortex and caudate-putamen. The distribution of immunoreactivity in the subcellular fraction were as follows: synaptic membranes > crude mitochondrial fraction > homogenate > myelin. In conclusion these studies suggest that: (a) the rat brain GBP and the bovine brain GBP are immunologically homologous protein, (b) there are no structural similarities between the GBP and the glutamate metabolizing enzymes with the exception of glutamate decarboxylase and (c) the subcellular and regional distribution of the GBP immunoreactivity followed a similar pattern as observed for L-(/sup 3/H)-binding.

• Research Organization: Kansas Univ., Lawrence (USA)
• OSTI ID: 6994783
• Country of Publication: United States
• Language: English

Related Subjects

550201* -- Biochemistry-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
AMINO ACIDS
ANIMAL TISSUES
ANIMALS
ANTIBODIES
ANTITHYROID DRUGS
BIOCHEMICAL REACTION KINETICS
BODY
BRAIN
CARBON-CARBON LYASES
CARBONIC ACID DERIVATIVES
CARBOXY-LYASES
CARBOXYLIC ACIDS
CATTLE
CELL CONSTITUENTS
CELL MEMBRANES
CENTRAL NERVOUS SYSTEM
CEREBELLUM
CEREBRAL CORTEX
CEREBRUM
DECARBOXYLASES
DISTRIBUTION
DOMESTIC ANIMALS
DRUGS
ENZYME IMMUNOASSAY
ENZYMES
GLUTAMIC ACID
HORMONE ANTAGONISTS
HYDROLASES
IMMUNE REACTIONS
IMMUNOASSAY
KINETICS
LABELLED COMPOUNDS
LIGASES
LYASES
MAMMALS
MEMBRANE TRANSPORT
MEMBRANES
MITOCHONDRIA
MYELIN
NERVE TISSUE
NERVOUS SYSTEM
ORGANIC ACIDS
ORGANIC COMPOUNDS
ORGANIC SULFUR COMPOUNDS
ORGANOIDS
ORGANS
OXIDOREDUCTASES
PEPTIDE HYDROLASES
PROTEINS
PURIFICATION
RATS
REACTION KINETICS
RODENTS
RUMINANTS
SUBCELLULAR DISTRIBUTION
THIOCYANATES
TISSUES
TRITIUM COMPOUNDS
VERTEBRATES