Purification of melanoma growth stimulatory activity
The Hs0294 human malignant melanoma cell line produces a monolayer mitogen that stimulates the serum free growth of low-density cultures of Hs0294 cells. This report describes the purification of that mitogen, termed MGSA for melanoma growth stimulatory activity, from serum-free conditioned medium from the Hs0294 cultures. MGSA has been purified from acetic acid extracts of lyophilized conditioned medium by gel filtration, reverse-phase high-pressure liquid chromatography (RP-HPLC), and preparative electrophoresis, resulting in a >400,000-fold purification. MGSA bioactivity resides in acid- and heat-stable polypeptides of high and low molecular weight. However, the majority of the activity is reproducibly associated with the approx.16-kd moiety eluting from RP-HPLC at approx.35% acetonitrile. Reduction with dithiothreitol or B-mercaptoethanol results in a loss of biological activity but does not convert the 24-28-kd moieties to the <14-16-kd forms of MGSA. /sup 125/I-MGSA that has been purified by preparative electrophoresis (16 kd) specifically binds to Hs0294 melanoma cells and retains 100% of the growth-stimulatory activity. The 16-kd MGSA stimulates the proliferation of Hs0294 cells at concentrations of 0.3-30 pM. Purified MGSA does not enable anchorage-independent growth of normal rat kidney (NRK) cells and is therefore different from the previously described transforming growth factors. The amino acid composition of MGSA differs from that of other previously described growth factors. These data demonstrate that MGSA represents a separate class of growth factors with biological and biochemical properties different from other growth factors.
- Research Organization:
- Emory Univ., Atlanta, GA
- OSTI ID:
- 6525409
- Journal Information:
- J. Cell. Physiol.; (United States), Vol. 129:3
- Country of Publication:
- United States
- Language:
- English
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MITOGENS
BIOCHEMISTRY
PURIFICATION
AMINO ACID SEQUENCE
ELECTROPHORESIS
GROWTH
IODINE 125
LIQUID COLUMN CHROMATOGRAPHY
MELANOMAS
POLYPEPTIDES
THYMIDINE
TRITIUM COMPOUNDS
TUMOR CELLS
ANIMAL CELLS
AZINES
BETA DECAY RADIOISOTOPES
CHEMISTRY
CHROMATOGRAPHY
DAYS LIVING RADIOISOTOPES
DISEASES
ELECTRON CAPTURE RADIOISOTOPES
HETEROCYCLIC COMPOUNDS
INTERMEDIATE MASS NUCLEI
IODINE ISOTOPES
ISOTOPES
LABELLED COMPOUNDS
MOLECULAR STRUCTURE
NEOPLASMS
NUCLEI
NUCLEOSIDES
NUCLEOTIDES
ODD-EVEN NUCLEI
ORGANIC COMPOUNDS
ORGANIC NITROGEN COMPOUNDS
PEPTIDES
PROTEINS
PYRIMIDINES
RADIOISOTOPES
RIBOSIDES
SEPARATION PROCESSES
550201* - Biochemistry- Tracer Techniques