EXPRESSION AND CHARACTERIZATION OF HUMAN LACTOFERRIN IN YEAST SACCHAROMYCES-CEREVISIAE

Lactoferrin (LF) has certain chemical and biological properties that a re significant to the dairy food industry. To study the relationship b etween protein structure and functionality of LF with the aim of incre asing the thermostability of LF, we established a yeast system for het erologous expression of cloned human LF cDNA. Human LF was successfull y synthesized in yeast cells by placing the cloned cDNA under the regu lation of yeast chelatin promoter. Both human LF and yeast invertase s ecretion signal sequences were used to direct the secretion of recombi nant human LF synthesized in yeast cells. The construct of the express ion unit containing the yeast invertase signal sequence resulted in re latively high levels of secretion of recombinant human LF (1.5-2.0 mg/ L), whereas the other containing human LF secretion signal sequence pr oduced quite low levels of secretion of the protein. The secreted reco mbinant human LF was purified from the yeast broth by heparin affinity and immunoaffinity chromatographies. The highly purified recombinant LF was confirmed to be bioactive, having iron-and copper-binding activ ities. Also, the recombinant LF synthesized in yeast was glycosylated. The levels of synthesis of human LF in the yeast system were not cons tant, and LF might be toxic to yeast cells.