Production of a 135-residue long N-truncated human keratinocyte growth factor 1 in Escherichia coli

Palifermin (trade name Kepivance®) is an amino-terminally truncated recombinant human keratinocyte growth factor 1 (KGF-1) with 140 residues that has been produced using Escherichia coli to prevent and treat oral mucositis following radiation or chemotherapy. In this study, an amino-terminally shortened KGF-1 variant with 135 residues was produced and purified in E. coli, and its cell proliferation activity was evaluated.

We expressed soluble KGF-1 fused to thioredoxin (TRX) in the cytoplasmic fraction of E. coli to improve its production yield. However, three N-truncated forms (KGF-1 with 140, 138, and 135 residues) were observed after the removal of the TRX protein from the fusion form by cleavage of the human enterokinase light chain C112S (hEK_L C112S). The shortest KGF-1 variant, with 135 residues, was expressed by fusion with TRX via the hEK_L cleavage site in E. coli and purified at high purity (> 99%). Circular dichroism spectroscopy shows that purified KGF-1₁₃₅ had a structure similar to that of the KGF-1₁₄₀ as a random coiled form, and MCF-7 cell proliferation assays demonstrate its biological activity.

We identified variations in N-terminus-truncated KGF-1 and selected the most stable form. Furthermore, by a simple two-step purification, highly purified KGF-1₁₃₅ was obtained that showed biological activity. These results demonstrate that KGF-1₁₃₅ may be considered an alternative protein to KGF-1.