Amino acid sequence of the mRNA cap-binding protein from human tissues
The 25-kDa mRNA cap-binding protein (CBP) involved in translation was purified by affinity chromatography from human erythrocytes and rabbit reticulocytes. The sequences of eight human and seven rabbit tryptic and V8 proteolytic peptides were determined. Based on the peptide sequence data, oligodeoxynucleotide probes were synthesized and used to screen human fibroblast and lymphocyte lambda cDNA libraries. The DNA sequence obtained from recombinant lambda phage inserts was found to code for all but one peptide. A 23-base oligonucleotide was synthesized based on the DNA sequence and used to prime synthesis of cDNA from human placental mRNA to construct a third library in lambdagt10. Screening with a 22-base oligonucleotide, whose sequence was upstream from the 23-base primer, yielded numerous recombinant phages with approx.250-base inserts. The 1900-base-pair cDNA sequence compiled from all phage inserts appeared to represent the entire primary sequence of CBP. Blot analysis of human placental and HeLa mRNA revealed multiple CBP mRNA species ranging from 1925 to 2250 bases. The amino acid sequence of CBP showed homology to the cap-binding PB2 protein of influenza virus.
- Research Organization:
- Univ. of Kentucky College of Medicine, Lexington
- OSTI ID:
- 5722852
- Journal Information:
- Proc. Natl. Acad. Sci. U.S.A.; (United States), Vol. 84:4, Issue 4
- Country of Publication:
- United States
- Language:
- English
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MESSENGER-RNA
MOLECULAR BIOLOGY
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AMINO ACID SEQUENCE
DNA SEQUENCING
ERYTHROCYTES
HELA CELLS
MAN
PHOSPHORUS 32
PLACENTA
RABBITS
RECOMBINANT DNA
RETICULOCYTES
ANIMALS
BETA DECAY RADIOISOTOPES
BETA-MINUS DECAY RADIOISOTOPES
BIOLOGICAL MATERIALS
BLOOD
BLOOD CELLS
BODY FLUIDS
DAYS LIVING RADIOISOTOPES
DNA
FETAL MEMBRANES
ISOTOPES
LIGHT NUCLEI
MAMMALS
MATERIALS
MEMBRANES
MOLECULAR STRUCTURE
NUCLEI
NUCLEIC ACIDS
ODD-ODD NUCLEI
ORGANIC COMPOUNDS
PHOSPHORUS ISOTOPES
PRIMATES
RADIOISOTOPES
RNA
STRUCTURAL CHEMICAL ANALYSIS
VERTEBRATES
550201* - Biochemistry- Tracer Techniques