The design of chimeric proteins is a major held of interest in structural biology and biotechnology. The successful design of the chimeric protein composed by the minimized reactive site domain of the low-molecular-mass trypsin inhibitor from Brassica napus (var. oleifera) seed (Ser3-Lys35; mini-RTI-III) and murine dihydrofolate reductase (DHFR) is reported here. The DHFR-mini-RTI-III chimeric protein was expressed in Escherichia coli, purified by metal-chelate affinity chromatography and oxidatively refolded. The affinity of the purified and refolded DHFR-mini-RTI-III for bovine trypsin (K = 5.0 x 10(-10) M) was closely similar to that determined for native RTI-III (K = 2.9 x 10(-10) M), at pH 8.2 and 22.0 degrees C. DHFR-mini-RTI-III may be regarded as a tool in structure-function studies and for developing multifunctional and multidomain proteinase inhibitors. (C) 2000 Academic Press.
Trovato, M., Maras, B., Polticelli, F., Costantino, P., Ascenzi, P. (2000). A chimeric mini-trypsin inhibitor derived from the oil rape proteinase inhibitor type III RID A-2020-2010 RID A-4573-2009. BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS, 275(3), 817-820 [10.1006/bbrc.2000.3376 WC Biochemistry & Molecular Biology; Biophysics].