Preparation of biologically active platelet-derived growth factor type BB from a fusion protein expressed in Escherichia coli

Journal Article · Tue Apr 04 04:00:00 EDT 1989 · Biochemistry; (USA)

DOI:https://doi.org/10.1021/bi00433a032· OSTI ID:5365418

Hoppe, J; Weich, H A; Eichner, W ^[1]

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1. GBF--Gesellschaft fuer Biotechnologische Forschung mbH, Braunschweig (West Germany)

Preparations of the mitogen platelet-derived growth factor (PDGF) from human platelets contain two related polypeptides termed A chain and B chain. PDGF-B is highly homologous to a portion of p28{sup v-sis}, the transforming protein of simian sarcoma virus. The authors have studied the mitogenic potential of a PDGF-BB-like homodimer by expressing the sequence coding for the mature part of PDGF-B in Escherichia coli. Expression was achieved as cro-{beta}-gal-PDGF-B fusion protein which was exclusively found in the inclusion bodies. A monomeric PDGF-B fragment shortened by 12 amino acid residues from the NH{sub 2} terminus was excised from the fusion protein by CNBr cleavage. After protection of thiols by S-sulfonation, this fragment was purified by gel permeation chromatography and reversed-phase high-performance liquid chromatography. This monomeric protein was dimerized in the presence of a mixture of reduced and oxidized glutathione to yield biologically active rPDGF-BB with an overall yield of {approx}0.7 mg of rPDGF-BB/L of culture. Escherichia coli rPDGF-BB stimulated ({sup 3}H)thymidine incorporation into AKR2B fibroblast at concentrations of about 1 ng/mL.

• OSTI ID: 5365418
• Journal Information: Biochemistry; (USA), Journal Name: Biochemistry; (USA) Vol. 28:7; ISSN 0006-2960; ISSN BICHA
• Country of Publication: United States
• Language: English

Related Subjects

550201* -- Biochemistry-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
AMINO ACID SEQUENCE
AZINES
BACTERIA
BIOLOGICAL FUNCTIONS
BIOLOGICAL MATERIALS
BLOOD
BLOOD CELLS
BLOOD PLATELETS
BODY FLUIDS
CELL CONSTITUENTS
CHROMATOGRAPHY
ESCHERICHIA COLI
FUNCTIONS
GROWTH FACTORS
HETEROCYCLIC COMPOUNDS
HYDROGEN COMPOUNDS
LIQUID COLUMN CHROMATOGRAPHY
MATERIALS
MICROORGANISMS
MITOGENS
MOLECULAR STRUCTURE
NUCLEOSIDES
NUCLEOTIDES
ORGANIC COMPOUNDS
ORGANIC NITROGEN COMPOUNDS
ORGANIC SULFUR COMPOUNDS
PLASMIDS
PROTEINS
PURIFICATION
PYRIMIDINES
RIBOSIDES
SEPARATION PROCESSES
THIOLS
THYMIDINE
TRITIUM COMPOUNDS