Identification and analysis of a novel protein-tyrosine kinase from bovine thymus
A cytosolic protein-tyrosine kinase has been identified and purified to near homogeneity from calf thymus by using the phosphorylation of the tyrosine-containing peptide angiotensin I as an assay. Specific peptide phosphorylating activity was enhanced by carrying out the assay at high ionic strength (2M NaCl). The inclusion of NaCl at this concentration acts to stimulate endogenous protein-tyrosine kinase activity while simultaneously inhibiting other endogenous kinases. The purification procedure involved extraction of the enzyme from calf-thymus and sequential chromatography on columns of DEAE-cellulose, heparin-agarose, casein-sepharose, butylagarose, and Sephadex G-75. Analysis of the most highly purified preparations by sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed a single Coomassie blue-stained band of 41 KDa. This molecular weight was consistent with results obtained from gel filtration, indicating that the enzyme exists as a monomer. The enzyme has also been found to catalyze an autophosphorylation reaction. Incubation of the enzyme with Mn/sup 2 +/ and (..gamma..-/sup 32/P)ATP led to its modification on a tyrosine residue. Phosphopeptide mapping experiments indicated that the 41 KDa kinase was distinct from p56, the major membrane-associated protein-tyrosine kinase in T lymphocytes.
- Research Organization:
- Purdue Univ., West Lafayette, IN
- OSTI ID:
- 7108485
- Report Number(s):
- CONF-8606151-; TRN: 86-039092
- Journal Information:
- Fed. Proc., Fed. Am. Soc. Exp. Biol.; (United States), Vol. 45:6; Conference: 76. annual meeting of the Federation of American Society for Experimental Biology, Washington, DC, USA, 8 Jun 1986
- Country of Publication:
- United States
- Language:
- English
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PHOSPHOTRANSFERASES
ENZYME ACTIVITY
PURIFICATION
ATP
CATTLE
CHROMATOGRAPHY
ELECTROPHORESIS
LYMPHOCYTES
MANGANESE COMPOUNDS
MOLECULAR WEIGHT
PHOSPHORUS 32
PHOSPHORYLATION
THYMUS
TRACER TECHNIQUES
ANIMAL CELLS
ANIMALS
BETA DECAY RADIOISOTOPES
BETA-MINUS DECAY RADIOISOTOPES
BIOLOGICAL MATERIALS
BLOOD
BLOOD CELLS
BODY
BODY FLUIDS
CHEMICAL REACTIONS
CONNECTIVE TISSUE CELLS
DAYS LIVING RADIOISOTOPES
DOMESTIC ANIMALS
ENZYMES
ISOTOPE APPLICATIONS
ISOTOPES
LEUKOCYTES
LIGHT NUCLEI
LYMPHATIC SYSTEM
MAMMALS
MATERIALS
NUCLEI
NUCLEOTIDES
ODD-ODD NUCLEI
ORGANIC COMPOUNDS
ORGANS
PHOSPHORUS ISOTOPES
PHOSPHORUS-GROUP TRANSFERASES
RADIOISOTOPES
RUMINANTS
SEPARATION PROCESSES
SOMATIC CELLS
TRANSFERASES
TRANSITION ELEMENT COMPOUNDS
VERTEBRATES
550201* - Biochemistry- Tracer Techniques