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Evidence for pyroglutamyl peptidase I and prolyl endopeptidase activities in the rat insulinoma cell line RINm 5F: lack of relationship with TRH metabolism
- Source :
- Molecular and Cellular Biochemistry, Molecular and Cellular Biochemistry, Springer Verlag, 1991, pp.15-24
- Publication Year :
- 1991
- Publisher :
- Springer Science and Business Media LLC, 1991.
-
Abstract
- International audience; Thyrotropin-Releasing hormone (TRH)-degrading pyroglutamyl peptidase I (PGP I) and prolylendopeptidase (PE) activities have been demonstrated in rat insulinoma RINm 5F cell line. These two enzymes catalyze the conversion of TRH to Histydyl-Proline-Diketopiperazine and to acid TRH respectively. After cell fractionation, we found all the PGP I and PE activities in the cytosolic fraction. The membrane-bound PGP II activity is not detectable in the RINm 5F cells. Further investigations on these two cytosolic enzymes show that pyroglutamyl- and proline-containing peptides are inhibitors of each TRH-degrading enzyme. Gel filtration chromatography on Sephadex G100 shows that PGP I and PE activity have an apparent molecular mass of about 18 kDa and 57 kDa, respectively. Kinetic analysis with TRH as substrate, gives a Km of 44 microM and 235 microM, and a Vmax of 1.49 and 8.80 pmol/min/micrograms protein for PGP I and PE, respectively. Immunoreactive TRH, His-Pro-Diketopiperazine and acid TRH levels in the cell line extracts are 2.2 +/- 0.9, 22.5 +/- 11.1 and 28.7 +/- 14.6 pg/1O6 cells, respectively. When cells have been incubated for 2 to 72 hours with a P.E. inhibitor (Z-Gly-Pro-CHN2) at 5 x 10(-7) M, both cell PGP I and PE activities are inhibited. No change in the cellular content of immunoreactive TRH, His-Pro-Diketopiperazine and acid TRH have been observed in treated cells.(ABSTRACT TRUNCATED AT 250 WORDS)
- Subjects :
- endocrine system diseases
Clinical Biochemistry
Thyrotropin-releasing hormone
MESH: Amino Acid Sequence
Substrate Specificity
MESH: Pyroglutamyl-Peptidase I
MESH: Animals
MESH: Serine Endopeptidases
MESH: Endopeptidases
Thyrotropin-Releasing Hormone
chemistry.chemical_classification
MESH: Protease Inhibitors
MESH: Kinetics
MESH: Molecular Weight
Serine Endopeptidases
General Medicine
Adenoma, Islet Cell
Biochemistry
Enzyme inhibitor
Cell fractionation
Prolyl Oligopeptidases
MESH: Diazomethane
hormones, hormone substitutes, and hormone antagonists
Subcellular Fractions
medicine.drug
endocrine system
MESH: Rats
Molecular Sequence Data
Radioimmunoassay
MESH: Adenoma, Islet Cell
Pyroglutamyl-Peptidase I
[SDV.CAN]Life Sciences [q-bio]/Cancer
Biology
Peptide hormone
Cell Line
MESH: Radioimmunoassay
[SDV.CAN] Life Sciences [q-bio]/Cancer
Prolyl endopeptidase
MESH: Thyrotropin-Releasing Hormone
Endopeptidases
medicine
Animals
Protease Inhibitors
Amino Acid Sequence
Pyroglutamyl-peptidase I
Molecular Biology
MESH: Molecular Sequence Data
Rat Insulinoma
Cell Biology
MESH: Cell Line
Rats
Molecular Weight
Kinetics
Enzyme
Diazomethane
chemistry
MESH: Subcellular Fractions
biology.protein
MESH: Substrate Specificity
Subjects
Details
- ISSN :
- 15734919 and 03008177
- Volume :
- 106
- Database :
- OpenAIRE
- Journal :
- Molecular and Cellular Biochemistry
- Accession number :
- edsair.doi.dedup.....93fe68c645ec8189adb00dfabf09a9df