A rapid and efficient expression and purification system has been developed for large-scale production of biologically active recombinant human keratinocyte growth factor-2 (rhKGF-2). The gene encoding human KGF-2 was cloned into the expression vector pET3c and transformed into Escherichia coli BL21(DE3)/pLys S. Under optimal conditions in a 30-l fermentor, the average bacterial yield and the average expression level of rhKGF-2 of three batches were up to 732 g and 32%, respectively. The recombinant protein was purified by cation exchange and heparin-affinity chromatography. One hundred and sixty five milligrams of pure rhKGF-2 was achieved per liter culture. A preliminary biochemical characterization of purified rhKGF-2 was performed by Western blotting and mitogenic activity analysis, and the results demonstrated that purified rhKGF-2 could react with anti-human KGF-2 antibody and stimulate the proliferation of HaCat cells.
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Chellaiah AT, McEwen DG, Werner S, Xu J, Ornitz DM (1994) Fibroblast growth factor receptor (FGFR) 3. Alternative splicing in immunoglobulin-like domain III creates a receptor highly specific for acidic FGF/FGF-1. J Biol Chem 269:11620–11627
Emoto H, Tagashira S, Mattei MG, Yamasaki M, Hashimoto G, Katsumata T, Negoro T, Nakatsuka M, Birnbaum D, Coulier F, Itoh N (1997) Structure and expression of human fibroblast growth factor-10. J Biol Chem 272:23191–23194
Han DS, Li F, Holt L, Connolly K, Hubert M, Miceli R, Okoye Z, Santiago G, Windle K, Wong E, Sartor RB (2000) Keratinocyte growth factor-2 (FGF-10) promotes healing of experimental small intestinal ulceration in rats. Am J Physiol Gastrointest Liver Physiol 279:G1011–1022
Igarashi M, Finch PW, Aaronson SA (1998) Characterization of recombinant human fibroblast growth factor (FGF)-10 reveals functional similarities with keratinocyte growth factor (FGF-7). J Biol Chem 273:13230–13235
Jimenez PA, Rampy MA (1999) Keratinocyte growth factor-2 accelerates wound healing in incisional wounds. J Surg Res 81:238–242
Luo Y, Cho HH, Jones RB, Jin C, McKeehan WL (2004) Improved production of recombinant fibroblast growth factor 7 (FGF7/KGF) from bacteria in high magnesium chloride. Protein Expr Purif 33:326–331
Miceli R, Hubert M, Santiago G, Yao DL, Coleman TA, Huddleston KA, Connolly K (1999) Efficacy of keratinocyte growth factor-2 in dextran sulfate sodium-induced murine colitis. J Pharmacol Exp Ther 290:464–471
Miki T, Bottaro DP, Fleming TP, Smith CL, Burgess WH, Chan AM, Aaronson SA (1992) Determination of ligand-binding specificity by alternative splicing: two distinct growth factor receptors encoded by a single gene. Proc Natl Acad Sci U S A 89:246–250
Ornitz DM, Leder P (1992) Ligand specificity and heparin dependence of fibroblast growth factor receptors 1 and 3. J Biol Chem 267:16305–16311
Partanen J, Makela TP, Eerola E, Korhonen J, Hirvonen H, Claesson-Welsh L, Alitalo K (1991) FGFR-4, a novel acidic fibroblast growth factor receptor with a distinct expression pattern. EMBO J 10:1347–1354
Smith PD, Polo M, Soler PM, McClintock JS, Maggi SP, Kim YJ, Ko F, Robson CM (2000) Efficacy of growth factors in the accelerated closure of interstices in explanted meshed human skin grafts. J Burn Care Rehabil 21(1 Pt 1):5–9
Xia YP, Zhao Y, Marcus J, Jimenez PA, Ruben SM, Moore PA, Khan F, Mustoe TA (1999) Effects of keratinocyte growth factor-2 (KGF-2) on wound healing in an ischaemia-impaired rabbit ear model and on scar formation. J Pathol 188:431–438
The work was supported by grants from the National High-Tech Research and Development Program of China (2007AA02Z110) and the Science Foundation of Zhejiang Province of China (No. Z205755).
Xiaoping Wu and Haishan Tian contributed equally to this work.
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Wu, X., Tian, H., Huang, Y. et al. Large-scale production of biologically active human keratinocyte growth factor-2. Appl Microbiol Biotechnol 82, 439–444 (2009). https://doi.org/10.1007/s00253-008-1782-x
- Human keratinocyte growth factor-2
- Large-scale production
- Mitogenic activity