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Biotechnology and Applied Biochemistry (2001) 33, (117–121) (Printed in Great Britain)

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Improved preparation and crystallization of 25 kDa human fibroblast growth factor-9

Nobuyuki Koyama*, Hiroaki Ohmae†, Shinji Tsuji†, Yoko Tanaka†, Tsutomu Kurokawa‡ and Osamu Nishimura§¹

*Discovery Research Laboratory II, Pharmaceutical Discovery Research Division, Takeda Chemical Industries, Ltd., Wadai-10, Tsukuba, Ibaraki 300–4293, Japan, †Discovery Research Laboratory IV, Pharmaceutical Discovery Research Division, Takeda Chemical Industries, Ltd., Yodogawa-ku, Osaka 532–8686, Japan, ‡Discovery Research Laboratory III, Pharmaceutical Discovery Research Division, Takeda Chemical Industries, Ltd., Yodogawa-ku, Osaka 532–8686, Japan, and §Pharmaceutical Discovery Research Division, Takeda Chemical Industries, Ltd., Wadai-10, Tsukuba, Ibaraki 300–4293, Japan

Abbreviations used: hFGF-9, human fibroblast growth factor-9; hFGF N33, 25 kDa hFGF-9; TFA, trifluoroacetic acid.

¹ To whom correspondence should be addressed (e-mail Nishimura_Osamu@takeda.co.jp).

We prepared 25 kDa human fibroblast growth factor-9 (hFGF-9 N33) on a large scale after overproduction in Escherichia coli MM294 (DE3)/pTG931. The purification was performed by a combination of hydrophobic chromatography and HPLC with an ion exchange column, a heparin affinity column and a gel filtration column. This improved procedure was rapid and simple, and the purified hFGF-9 N33 was found to be homogeneous as judged by various criteria, such as amino acid analysis, N-terminal amino acid sequence, C-terminal amino acid analysis and biological activity. Furthermore, as determined by low endotoxin and DNA content, the protein was of high purity. In addition, the hFGF-9 N33 prepared in the present study was easily crystallized by the vapour diffusion method.

Received 26 September 2000; accepted 25 November 2000

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