Characterization and biological activities of recombinant human plasminogen kringle 1-3 produced in Escherichia coli.
Angiogenesis, the formation of new capillaries from preexisting blood vessels, is involved in many pathological conditions, for example, tumorigenesis, diabetic retinopathy, and rheumatoid arthritis. Angiostatin, which contains the kringle 1-4 domains of plasminogen, is known to be a potent inhibitor of angiogenesis and a strong suppressor of various solid tumors. In this study, we expressed recombinant protein containing the kringle 1-3 domains of human plasminogen in Escherichia coli and investigated its biological activities. The protein was successfully refolded from inclusion bodies and purified at a 30% overall yield, as a single peak by HPLC. The purified recombinant protein had biochemical properties that were similar to those of the native form, which included molecular size, lysine-binding capacity, and immunoreactivity with a specific antibody. The recombinant protein was also found to strongly inhibit the proliferation of bovine capillary endothelial cells in vitro, and the formation of new capillaries on chick embryos. In addition, it suppressed the growth of primary Lewis lung carcinoma and B16 melanoma in an in vivo mouse model. Our findings suggest that the recombinant kringle 1-3 domains in a prokaryote expression system have anti-angiogenic activities, which may be useful in clinical and basic research in the field of angiogenesis.
You
Weon-Kyoo
WK
Mogam Biotechnology Research Institute, Bioproducts Research Center, 341 Pojung-ri, Koosung-myun, Yongin City, Kyonggi-do 449-910, Republic of Korea.
So
Seung-Ho
SH
Sohn
Young-Doug
YD
Lee
Hyosil
H
Park
Doo-Hong
DH
Chung
Soo-Il
SI
Chung
Kwang-Hoe
KH
D000595:Amino Acid Sequence; D020533:Angiogenesis Inhibitors / Q000737:chemistry* / Q000302:isolation & purification / Q000494:pharmacology*; D043168:Angiostatins / Q000494:pharmacology; D000818:Animals; D000970:Antineoplastic Agents / Q000737:chemistry* / Q000302:isolation & purification / Q000494:pharmacology*; D018827:Carcinoma, Lewis Lung / Q000378:metabolism; D002417:Cattle; D002465:Cell Movement / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D042783:Endothelial Cells / Q000187:drug effects; D004926:Escherichia coli / Q000235:genetics*; D016222:Fibroblast Growth Factor 2 / Q000037:antagonists & inhibitors; D006801:Humans; D051379:Mice; D008810:Mice, Inbred C57BL; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010446:Peptide Fragments / Q000737:chemistry* / Q000235:genetics / Q000494:pharmacology*; D010958:Plasminogen / Q000737:chemistry* / Q000235:genetics / Q000494:pharmacology*; D017510:Protein Folding; D023041:Xenograft Model Antitumor Assays
D020533:Angiogenesis Inhibitors; D000970:Antineoplastic Agents; D010446:Peptide Fragments; C402400:recombinant kringle 1-3 (rKI-3) of human plasminogen; D016222:Fibroblast Growth Factor 2; D043168:Angiostatins; D010958:Plasminogen