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Novel bombesin-like peptide binding proteins from lung
M W Geraci1, Y E Miller, A Escobedo-Morse
1Division of Pulmonary Sciences, University of Colorado Health Sciences Center, Denver.
Summary
Researchers identified novel gastrin-releasing peptide (GRP) binding proteins in mouse lung cytoplasm. These proteins exhibit high-affinity GRP binding and were characterized using affinity chromatography and sequencing, revealing unique peptide sequences.
Area of Science:
- Biochemistry
- Molecular Biology
- Pulmonology
Background:
- Gastrin-releasing peptide (GRP) and bombesin-like peptides (BLP) are crucial in lung development, injury response, and cancer.
- Existing BLP receptor mRNAs are undetectable in normal lung tissue via Northern blot.
Purpose of the Study:
- To isolate and characterize BLP binding proteins from normal mouse lung.
- To investigate the presence and nature of GRP binding sites beyond known receptors.
Main Methods:
- Preparation of soluble cytoplasmic and detergent-solubilized membrane fractions from mouse lung.
- Assessment of specific 125I-GRP binding using saturation assays.
- Isolation of BLP binding proteins via GRP14-27 affinity chromatography, SDS-PAGE, HPLC, trypsin digestion, and Edman degradation sequencing.
Main Results:
- Both soluble cytoplasmic and membrane fractions showed saturable, high-affinity GRP binding (Kd = 1.6 nM and 7.5 nM, respectively).
- Affinity chromatography yielded a prominent 70 kD band, with weaker 65 and 75 kD bands, in both fractions.
- Sequencing of cytoplasmic BLP binding proteins revealed unique peptide sequences not found in databases.
Conclusions:
- Normal mouse lung contains previously unidentified, high-affinity GRP binding proteins in both cytoplasmic and membrane fractions.
- These novel proteins, particularly the 70 kD cytoplasmic form, are distinct from known BLP receptors.
- The identified unique peptide sequences provide a basis for further structural and functional characterization of these novel lung BLP binding proteins.