Full-length recombinant human SCF1-165 is more thermostable than the truncated SCF1-141 form

Abstract

Human stem cell factor initiates a diverse array of cellular responses, including hematopoiesis, cell proliferation, differentiation, migration and survival. To explore the relationship between its structure and function, we produced recombinant soluble human stem cell factor1-165 (wild type) and human stem cell factor1-141 (C-terminal truncated) in a yeast expression system and compared their biological activities and thermal stabilities. The biological activity of the two proteins was measured as a function of TF-1 cell viability and effects on downstream signaling targets after incubation. We found that these proteins enhanced cell viability and downstream signaling to a similar extent, in a dose-dependent manner. The biological activity of recombinant human stem cell factor1-165 was significantly greater than that of recombinant human stem cell factor 1-141 after heating the proteins (100 ng/mL) at 25-110°C for 10 minutes (P<0.05 for all temperatures). In addition, circular dichroism spectral analysis indicated that β-sheet structures were altered in recombinant human stem cell factor1-141 but not recombinant human stem cell factor1-165 after heating at 90°C for 15 or 30 min. Molecular modeling and limited proteolytic digestion were also used to compare the thermo stability between human stem cell factor1-165 and human stem cell factor1-141. Together, these data indicate that stem cell factor1-165 is more thermostable than stem cell factor 1-141. © 2014 Weng et al.